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i::!:;<ii.ili:illtlllli!:li:!ll'!ilillii!!l|!lii!iill!it;ii!l!H!H^ 


Chemical  Pathology 


BEING  A  DISCUSSION  OF  GENERAL  PATH- 
OLOGY FROM  THE  STANDPOINT  OF 
THE    CHEMICAL    PROCESSES     INVOLVED 


BY 

H.  GIDEON  WELLS,  Ph.D.,  M.D. 

PROFESSOR    OF    I'ATIIOLOGY    IN    THE    UNIVERSITY    OF    CHICAGO    ANU    IN 

RUSH    MEDICAL   COLLEGE.    CHICAGO;    DIRECTOR    OP   THE 

OTHQ   S.    A.    SPRAGOE    MEMORIAL    INSTITUTE 


TJIIRD  E  Din  ON,  REVISED  AND  RESET 


PHILADELPHIA  AND  LONDON 


W.    B.    SAUNDERS    COMPANY 

1918 


x:5 


*^^' 


Copyright,     1907,    by    VV.     B.     Saunders    Company.     Revised,    entirely 
reset,    reprinted    and    recopyrighted,    February.     1014.     Revised,    entirely 
reset,   reprinted  and   recopyrighted,  January,    igiti. 

Copyright.    19 18.    by    W.    B.    Saunders    Company 


PRINri;i>     l.\     AMERUA 


TO 

XuOvici   Ibelitoen 

THIS  BOOK  IS  RESPECTFULLY  DEDICATED.  AS  A 

SLIGHT  TOKEN  OF  THE  GRATITUDE  AND 

ESTEEM  OF  HIS  PUPIL 


nSOBOl 


PREFACE  TO  THE  THIRD  EDITION 

Despite  the  war,  active  investigations  in  the  chemical  problems  of 
disease  have  continued,  even  in  those  countries  most  deeply  involved 
in  the  conflict.  Although  some  of  the  later  publications  of  foreign 
countries  have  not  been  directly  accessible,  but  few  have  not  been  avail- 
able at  least  through  abstracts,  and  it  is  believed  that  most  of  the 
literature  of  importance,  within  the  scope  of  this  book,  has  been  con- 
sidered in  its  revision,  although  the  rule  previously  followed  of  quot- 
ing only  from  the  original  articles  has  of  necessity  been  violated  in 
several  instances.  The  new  additions  to  our  knowledge  in  the  three 
years  since  the  second  edition  was  issued  have  been  so  numerous  that 
it  has  again  been  necessary  to  reprint  the  entire  work.  Several  sub- 
jects have  been  largely  rewritten,  especially  Gout,  Specificity  of  Im- 
munological Reactions,  Anaphylaxis,  Icterus,  Acidosis,  Diabetes  and 
Uremia.  New  sections  have  been  added  on  the  Abderhalden  Reaction, 
Specificity,  Chemical  Basis  of  Growth,  Atrophy,  and  the  Pressor 
Bases,  as  well  as  many  briefer  additions.  As  previously,  every  effort 
has  been  made  to  make  the  discussion  of  each  topic  as  brief  as  con- 
sistent with  reasonable  clearness  and  completeness.  Where  new 
articles  including  older  references  have  been  quoted,  the  latter  have, 
in  most  instances,  been  omitted  from  the  book. 

Again  I  must  express  my  indebtedness  to  the  several  colleagues 
who  have  been  so  kind  as  to  read  over  various  sections  of  the  book  and 
to  help  me  with  their  suggestions;  and  also  to  the  members  of  my 
Department  who  have  helped  me  so  generously  with  the  proof  reading, 
especially  Dr.  Lydia  ]\I.  De  Witt  and  Miss  Harriet  F.  Holmes. 

Dr.  R.  T.  Woodyatt  has  kindly  revised  the  chapter  on  Diabetes, 
which  he  contributed  to  the  second  edition.  Since  that  edition  was 
printed  much  valuable  information  on  the  subject  of  carbohydrate 
metabolism  has  been  contributed  from  Dr.  Woodyatt 's  own  labora- 
tory, and  through  his  new  method  for  accurately  timed  and  measured 
intravenous  injections  the  way  has  been  opened  for  many  advances  in 
the  study  of  metabolism  under  both  normal  and  pathological  condi- 
tions. 

H.  G.  W. 

Chicago,  III.,  November,  1917. 


PREFACE  TO  THE  FIRST  EDITION 

DxjKiNG  the  past  score  of  years  the  subject  of  biological  chemistry 
has  attracted  the  attention  and  labors  of  a  constantly  increasing  num- 
ber of  investigators,  many  of  whom  have,  for  one  reason  or  another, 
been  interested  in  ])atlu)logical  conditions.  Sometimes  the  physiologist 
has  souglit  for  light  on  his  problems  in  the  evidence  afforded  by  related 
pathological  conditions.  Frequently  clinicians  have  studied  the  meta- 
bolic clianges  and  the  composition  of  the  products  of  disease  processes. 
Relatively  seldom,  unfortunately,  has  the  pathologist  attacked  his 
problems  by  chemical  methods.  From  the  above  and  other  sources 
have  come  scattered  fragments  of  information  concerning  the  chemi- 
cal changes  that  occur  in  pathological  phenomena.  Only  when  bearing 
upon  conditions  such  as  gout  and  diabetes,  which  concern  alike  the 
physiologist,  the  clinician,  and  the  pathologist,  have  the  fragments 
been  moulded  together  into  a  homogeneous  whole.  For  the  most  part 
they  still  remain  isolated,  uncorrelated,  frequently  unconfirmed  items 
of  information,  scattered  through  medical,  chemical,  physiological,  and 
physical  literature. 

It  has  been  the  aim  of  the  writer  to  collect  these  scattered  fragments 
as  completely  as  possible,  and  to  use  them  as  a  basis  for  a  consideration 
of  General  Pathology  from  the  standpoint  of  the  chemical  processes 
which  occur  in  pathological  conditions.  Owing  to  the  diffuselj^  scat- 
tered conditions  of  the  literature  on  which  this  work  is  based,  it  cannot 
be  claimed  that  all  of  the  many  contributions  from  which  useful  in- 
formation might  be  obtained  have  been  noticed ;  but  it  is  hoped  that 
a  sufficiently  thorough  collection  of  material  has  been  made  to  afford 
a  fair  basis  for  a  consideration  of  "Chemical  Pathology."  The  time 
seems  ripe  for  an  effort  of  this  nature.  Within  the  past  few  years 
great  and  encouraging  advances  have  been  made  in  biological  chem- 
istry, which  in  many  instances  seem  to  throw  light  upon  pathological 
processes.  In  medicine,  the  use  of  chemical  methods  in  the  study  of 
clinical  manifestations  has  become  more  general,  and  has  yielded 
valuable  information.  Pathologists  have  come  to  feel  that  the  op- 
portunities for  the  acquirement  of  knowledge  by  means  of  morphologi- 
cal studies  have  become  reduced  to  a  mininmm,  while  the  fields  of 
pathological  phj^siology  and  chemistry  lie  still  almost  unexplored. 
The  development  of  research  upon  the  subject  of  natural  and  acquired 
immunity  has  presented  innumerable  problems,  all  of  which  are 
essentialiv  chemical.     And  perhaps  most  important  of  all  is  the  general 


8  P  KEF  ACE  TO  THE  FIRST  EDITION 

av:akonIn^  of  an  appreciation  of  the  importance  of  physiological  chem- 
it.tr>  to  medical  science,  wtiich  has  led  to  the  introduction  of  laboratory 
courses  on  this  subject  in  every  medical  school  worthy  of  the  name. 

A  book  on  Chemical  Pathology  should,  therefore,  seek  to  supply 
information  to  a  varied  group  of  readers.  It  sliould  furnish  collateral 
reading  to  the  student  who  for  the  first  time  goes  over  the  subject  of 
General  Pathology,  which  his  text-books  usually  consider  chiefly  from 
the  morphological  standpoint.  It  should  exploit  to  the  graduate  in 
medicine  tlie  advances  that  are  being  made  along  lines  that  are  of 
fundamental  importance  to  clinical  medicine.  It  should  serve  for  the 
investigator  in  biological  chemistry  or  in  pathology  as  a  source  of 
information  concerning  the  ground  upon  which  the  two  subjects  over- 
lap— the  "Grenzgebiete"  of  Pathology  and  Physiological  Chemistry. 
And,  above  all,  it  should  afford  a  guide  to  the  sources  of  our  knowledge 
of  these  subjects,  since  nothing  but  direct  familiarity  with  the  original 
reports  of  the  investigators  themselves  can  give  the  student  an  im- 
personal view  of  the  actual  status  of  the  questions  under  consideration. 
On  account  of  this  multiplicity  of  the  objects  in  view,  it  has  often  been 
necessary  to  consider  certain  topics  from  more  than  one  standpoint ; 
which  explains,  perhaps,  certain  apparent  irregularities  in  the  style 
and  manner  of  treatment. 

It  has  been  assumed  that  the  reader  has  at  least  an  elementary 
knowledge  of  organic  and  physiological  chemistry.  For  the  benefit 
of  those  whose  studies  in  these  subjects  date  back  some  years,  it  has 
seemed  advisable  to  include  in  an  introductory  chapter  an  epitome  of 
the  more  modern  views  concerning  the  chemistry  of  the  protein  mole- 
cule, the  composition  of  the  animal  cell,  and  the  principles  of  physical 
chemistry,  in  as  far  as  they  apply  to  biological  problems.  The  general 
consideration  of  ''Enzymes"  in  Chapter  II  is  written  with  a  similar 
object.  In  discussing  these  fundamental  topics  it  has  seemed  advis- 
able to  omit  detailed  references  to  the  numerous  original  sources, — 
these  may  be  found  quoted  in  the  special  text-books  cited  in  the  foot- 
notes; but  in  presenting  the  more  distinctly  pathological  topics  the 
attempt  has  been  made  to  render  all  the  important  literature  available 
to  the  reader  and  investigator.  To  economize  space,  a  complete  bibli- 
ography has  not  been  inserted  when  this  exists  already  eollected  in 
some  readily  accessible  review  or  original  article ;  hence  the  references 
cited  in  the  foot-notes  will  generally  be  found  to  include  only  the  more 
recent  publications.  These  references  have  been  so  selected,  however, 
that  they  will  be  found  to  furnish  bibliogra]ihical  matter  sufficient  to 
lead  the  investigator  to  all  the  important  litei-ature  on  the  topics 
covered  in  this  book.  As  to  those  subjects  (such  as  gout,  diabetes,  and 
gastro-intestinal  putrefaction)  which,  because  of  their  great  practical 
clinical  interest,  have  already  been  discussed  in  available  monographs 
at  greater  length  than  the  scope  of  this  work  would  })ermit,  it  has 
seemed  appropriate  merely  to  summarize  the  most  recent  views  and 


PREFACE  TO  THE  FIRST  EOmON  9 

advances,  referring  the  reader  to  tlie  ,s])eeial  treatises  for  the  general 
and  historical  discussions. 

It  is  with  the  greatest  pleasure  that  I  acknowledge  my  indebtedness 
to  many  colleagues  in  the  University  of  Chicago,  who  have  kindly  read 
the  sections  of  my  manuscript  tliat  touch  upon  tlieir  own  special  fields, 
and  whose  criticism  and  advice  have  been  of  the  greatest  assistance ; 
their  number  alone  prevents  my  thanking  them  by  name.  Most  par- 
ticularly, however,  must  I  express  my  debt  to  my  former  instructor, 
Professor  Lafayette  B.  jMendel,  of  Yale  University,  whose  kindly 
criticism  and  suggestions  have  been  of  inestimable  value.  For  con- 
stant assistance  in  the  preparation  of  the  manuscript,  and  for  the 
revision  of  the  bibliography,  I  am  indebted  to  my  wife. 

H.  G.  W. 


CONTENTS 

CHAPTEU  i  I'AGK 

iMKODUCnoN 17 

The    ClIEMlSTKY    and    rilYSlCS    OF    THE    CELL 17 

Chemistry  of  the  Essential  Cell  Constituents 18 

Proteins It) 

Fats  and  Lipoids   (Lipins) 23 

Carboliydiates         25 

Inorganic   iSubstances 26 

The  Physical  Chemistry  of  the  Cell  and  Its  Constituents 26 

Crystalloids    and    iheir    Properties 27 

Colloids 34 

The  (Structure  of  the  Cell  in  Relation  to  Its  Chemistry  and  Physics      .      .  43 

The    iS'ucleus 44 

The   Cytoplasm 46 

The   Cell-wall 49 

CHAPTER  II 

Enzymes 53 

The  Nature  of  Enzymes  and  Their  Actions 54 

The    Principles    of    Enzyme    Action 56 

The    Toxicity    of    Enzymes 61 

Anti-enzymes 63 

The  Intracellular  Enzymes 68 

Oxidizing    Enzymes 68 

Lipase 77 

Amylase SO 

CHAPTER  III 

Enzymes   (Coniinued)         81 

Intracellular   Proteases    (Proteolytic  Enzymes),   Including  a  Considera- 
tion  of   Autolysis 81 

Autolysis 82 

Relation  of  Autolysis  to  Metabolism 87 

Defense  of  the  Cells  Against  Their  Autolytic  Enzymes 88 

Autolysis   in   Pathological   Processes 90 

CHAPTER  IV 

The    Chemistry    of    Bacteria    and    Their    Products 106 

Structure  and  Physical  Properties lOti 

Chemical   Composition 107 

Bacterial    Enzymes 113 

Poisonous  Bacterial  Products 120 

Ptomains 120 

Toxins 125 

Endotoxins 129 

Poisonous  Bacterial  Proteins 13] 

Bacterial    Pigments 132 

11 


12  CONtENTS 

CHAPTER  V  PAGE 

Chemistry  of  the  Animal  Parasites 134 

Protozoa         135 

Cestodes         137 

Nematodes 140 


CHAPTER  VI 

Phytotoxins  and  Zootoxins 144 

Phytotoxins         144 

Tlie    Toxin    Causing    Hay-fever 147 

Zootoxins 148 

Snake    Venoms 148 

Scorpion   Poison 157 

Spider  Poison 158 

Centipedes         159 

Bee   Poison 160 

Poisons  of  Dermal  Glands  of  Toads  and  Salamanders 161 

Poisonous    Fish 162 

Eel  Serum 164 

CHAPTER  VII 

Chemistry   of    the    Immunity    Reactions — Antigens,    Specificity,    Anti- 
toxins, Agglutinins,  Precipitins,  Anaphylaxis  or  Allergy,  Ab- 

derhalden  Reaction,  Opsonins,  and  Related  Subjects  ....  165 

Antigens         166 

Non-Protein    Antigens 167 

Specificity    of    Immune    Reactions 171 

Toxins    and   Antitoxins 177 

Chemical  Nature  of  Antitoxins 180 

Agglutinins  and  Agglutination 183 

Precipitins 189 

Anaphylaxis    or    Allergy 193 

The  Abderhalden  Reaction 204 

Opsonins 207 

Tlie   Meiostagtiiin   Reaction 208 

The  Epiphanin  Reaction 209 

CHAPTER  VIII 

Chemistry    of    the    Immunity    Reactions     (Continued) — Bacteriolysis, 

Hemolysis,  Complement  Fixation,  and  Serum  Cytotoxins       .      .  210 

Scrum    Bacteriolysis 210 

Cytotoxins 214 

Hemolysis   or   Erythrocytolysis 215 

Hemolysis  liy  Known  Chemical  and  Physical  Agencies 215 

Hemolysis  by  Serum 218 

Hemolysis   by    ]5acteria 224 

Hemolysis  by  Vegetable  Poisons ', 225 

Hemolysis  1)y  Venoms 228 

Hemolysis  in  Disease 229 

Complement   Fixation   and  Wassermann   Reactions 234 

Cytolysis   in   General 238 

CHAPTER  IX 

Chemical  Means  of  Defense  Against  Non-Antigenic  Poisons  ....  243 

Inorganic  Poisons 246 

Organic  Poisons 248 


CONTENTS  13 

CHAPTER  X  PAGE 

Inflammation,   Rkcknkkation,  GIrowth 253 

Ameboid  Motion  and  Phagocytosis 254 

Chemotaxis 254 

Chemotaxis   of   Leucocytes 256 

Phagocytosis 262 

Theories  of  Chemotaxis  and  Phagocytosis 266 

Artificial   Imitations  of  Ameboid  Movement    .      .      .• 267 

Relation  of  the  Above  Experiments  to  tlie  Plienomena  Exhibited  by  Leu- 
cocytes in   Inflammation 271 

Suppuration 276 

Composition  of  Pus 277 

Sputum 280 

Proliferation  and  Regeneration 283 

Growth  and   Repair.     "'Vitamines." 285 


CHAPTER  XI 

Disturbances  of  Circut>ation  and  Diseases  of  the  Blood 289 

The  Composition  of  the  Blood 289 

Hemorrhage         293 

Hemophilia 297 

Anemia  and  the  Specific  Anemias 300 

Secondary    Anemias 300 

Chlorosis 302 

Pernicious    Anemia 305 

Leukemia 307 

Hyperemia 312 

Active  Hyperemia 312 

Passive    Hyperemia 313 

Thrombosis 315 

Fibrin   Formation 315 

The  Formation  of  Thrombi 322 

Embolism 325 

Infarction 327 


CHAPTER  XII 

Edema 330 

Formation    of    Lymph 331 

Absorption  of  Lymph 338 

The  Causes  of  Edema 339 

Special    Causes   of    Edema 348 

Composition  of  Edematous  Fluids 352 

Varieties   of   Edematous    Fluids 359 

Chemistry  of  Pneumothorax 365 


CHAPTER  XIII 

Retrogressive  Changes    (Necrosis,   Gangrene,  Rigor  Mortis,   Parenchy- 
matous Degeneration) 367 

Necrosis 36/ 

Causes  of  Necrosis 3/1 

Varieties  of  Necrosis 381 

Fat   Necrosis 384 

Gangrene 388 

Rigor  Mortis 390 

Atrophy 393 

Cloudv    Swelling 394 


14  CONTENTS 

CHAPTER  XIV  PAGE 

Retrogressive  Processes  (Continned),  Fatty,  Amyloid,  Hyaline,  Colloid. 

AND   GLYCOGf:XIC   INFILTRATION   AND  DEGENERATION 397 

Fatty  Metamorpliosis         397 

Physiological   Formation  of  Fat 397 

Pathological    Fat   Accunuilation 399 

Causes    of    Fatty    .Metamorphosis          406 

Processes   Related   to    Fatty   Metamorphosis 410 

Adipocere 410 

Lipemia 412 

Pathological  Occurrence  of  Fatty  Acids 414 

Pathological  Occurrence  of  Cholesterol 415 

Amyloid    Degeneration 417 

The  Origin  of  Amyloid 421 

Hyaline   Degeneration 423 

Colloid    Degeneration 425 

Mucoid  Degeneration 427 

Glycogen    in    Pathological   Processes 428 

Physiological  Occurrence 429 

Pathological    Occurrence 431 

CHAPTER  XV 

Calcification,   Concretions,   and   Incrustations 435 

Calcification 435 

Occurrence  of   Pathological   Calcification 438 

Chemistry  of  the  Process  of  Calcification 439 

Osteomalacia 443 

Rickets 445 

Concretions 447 

Biliary    Calculi 448 

Urinary    Calculi  454 

Corpora    Amylacea 400 

Other    Less    Common    Concretions 461 

Pncumonokoniosis 465 

CHAPTER  XVI  I 

Pathological  Pigmentation 467 

Melanin 467 

Lipochrome 474 

Blood  Pigments 476 

Icterus 484 

Congenital  Hemolytic  Icterus 489 


CIL\PTER  X^^I 

The  Chemistry  of  Tumors 492 

A.  Chemistry  of  Tumors  in  General 404 

B.  (licniistry  of  Certain  Specific  Tumors 50!) 

(1  I    Benign  Tumors 50!) 

(2)    Malignant  Tumors 515 

Multiple  Myelomas  and  ^lyclopatliic  "Alliuniosuria" 51S 

CHAPTER  XVIII 

Pathological   Conditions   Due   to,   or   Associated   with.   Abnormalities 

IN  Metabolism,  Including  Autointoxication 523 

I'remia 525 

Toxemias  of    Pregnancy 533 


COyTEMS  15 

PAGE 

Eclampsia 533 

Acute  Yellow  Atrophy  of  the  Liver                   53<l 

C'lieniiccil  Llumges  of  Acute  Yellow  Atrophy 54:i 

Acid   Intoxication .  ;j.}7 

Diabetic    Coma 5.30 

Acid  Intoxication  in  Conditions  Other  Than  Diabetes 5.")7 

Fatigue 50 1 

The    roisons    Produced    in    Superficial    Burns 5(;2 


CHAPTER  XIX 

Gastro-Inti:stinal  '•Autoixtoxication'"  axd  PvElated  Metabolic  Disturb- 

AXCES oGG 

I.     The  Constituents  of  the  Digestive  Fluids     ....  567 

II.     Products  of  Normal  Digestion 5t;8 

III.     Products  of  Putrefaction  and  Fermentation 57n 

A.  Protein  Putrefaction 571 

The  Pressor   Bases 57t) 

Alkaptonuria 577 

Cystine  and  Cystinuria 582 

B.  Products  of  Fermentation  of  Carbohydrates 58.3 

C.  Products  of  the  Decomposition  of  Fats 584 

Results  of  Gastro-intestinal  Intoxication 585 

Acute  Intestinal  Obstruction 588 


CHAPTER  XX 

Chemical  Pathology  of  the  Ductless  Glaxds 590 

Diseases  of  the  Thyroid 500 

The  Functions  of  the  Thyroid   .      .             590 

Chemistry    of    the    Thyroid 593 

The  Parathyroids         597 

Chemistry  of  Goiter 599 

Myxedema  and  Cretinism 601 

Exophthalmic  Goiter         604 

The  Adrenals  and  Addison's  Disease 60S 

Addison's  Disease 613 

The  Hypophysis  and  Acromegaly 614 

Thymus  and  Other  Ductless  Glands 616 


CHAPTER  XXI 

URic-Acin    IMetaiiolism     axd    Gout 618 

The   Chemistry   of   Uric   Acid 618 

Formation    of    Uric    Acid 621 

Destruction    of    Uric    Acid 625 

The  Occurrence  of  Uric  Acid  in  the  Blood.  Tissues  and  Urine      .  .  626 

Gout         628 

Uric-acid  Infarcts         633 


CHAPTER  XXII 

Diabetes 635 

Carbohydrate    Phvsiologv       .  637 

The  Blood  Sugar  ".      .      ' 641 

The  State  of  the  Sugar  in  the  Blood 643 

Diose         645 

Trioses 645 


16  CONTENTS 

?AGE 

Tetroses         ...  048 

Pentoses         G49 

Chronic  Pentosuria 650 

Hexoses 650 

Galactose 654 

Levulose   (Fructose) 655 

Polysaccharides 656 

Glycosurias 657 

Phlorhizin  Diabetes 65!) 

Pancreas  Diabetes  and  Diabetes  Melitus 665 


IXDEX         073 


chp:]\iical  pathology 

CHAPTER    I 

INTRODUCTION 

THE  CHEMISTRY  AND  PHYSICS  OF  THE  CELL 

Since  Virchow  founded  modem  pathology  the  unit  of  all  anatomical 
considerations  of  disease  has  been  the  cell,  and  in  physiology  the  same 
unit  has  been  found  equally  useful.  When  either  physiological  or 
pathological  processes  are  studied  from  a  chemical  standpoint,  the 
cell  is  still  found  occupying  nearly  as  fundamental  a  position,  for 
we  can  seldom  go  back  to  molecules  and  atoms  in  investigating 
biological  problems.  Although  we  know  that  within  each  cell  are 
many  different  chemical  substances,  and  that  numerous  different 
enzj^mes  and  other  agencies  are  exerting  their  influences  upon  them, 
yet  we  find  that  the  reactions  are  all  profoundly  affected  by  the 
environment  in  which  they  occur,  and  it  is  the  stracture  of  the  cell 
that  determines  the  environment  of  its  chemical  constituents.  All 
chemical  reactions  are  modified  by  physical  influences,  and  an  enzyme 
may  have  quite  a  different  effect  upon  a  substance  when  it  acts  in  a 
test-tube  from  wliat  it  will  have  when  in  a  living  cell,  whose  struc- 
ture permits  the  diffusion  of  one  substance  while  preventing  that  of 
another,  and  where  countless  other  substances  and  enzymes  may 
participate  in  the  changes.  The  cell  is  the  structural  unit  of  the  liv- 
ing organism,  and  as  by  its  physical  properties  it  modifies  chemical 
processes,  so  it  becomes  practically  the  unit  in  physiological  and 
pathological  chemistry.  All  consideration  of  the  chemistry  of  disease 
must  thus  refer  back  to  the  chemistry  and  physics  of  the  normal  cell, 
and  on  this  account  a  brief  resume  of  these  subjects  may  serve  as  a 
fitting  introduction  to  the  strictly  pathological  matters  to  follow.^ 

As  a])plied  to  the  animal  tissues,  the  tenn  "cell"  is  entirely  a  mis- 
nomer, for  it  describes  accurately  only  such  forms  of  "cells"  as  are 

1  Of  necessity,  only  so  much  of  the  vorv  extensive  literature  on  coll  structure 
and  cell  chemistry  can  be  considered  as  will  have  direct  bearing  \ipon  tiie  subject 
matter  to  follow,  referring  tlie  reader  for  more  detailed  information  to  sucli  works 
as  Wilson's  "The  Cell  in  Development  and  Inlioritance";  Mann's  "Physioloirical 
Histoloffy"';  Hammarsten's  "Physiolofrical  Cliemistry":  Gurwitscli's  "Morplioloeie 
imd  Biolo.srie  der  Zelle";  Hober's  "Pliysikalisclie  Cbemie  der  Zelle  luid  der 
Clewebe";  Hamburojer's  "Osmotischer  Druck  und  Tonenlehre";  Loeb's  "nviiamics 
of  Living  Matter";  Oppenheimer's  "ITandbuch  der  Biochemie";  and  Botta-'zi, 
"Handbuch  der  vergl.  Pliysiologie,"  Vol.  T,  for  general  discussion,  and  to  the  most 
important  monographs  for  treatment  of  special  points. 
2  17 


18  THE    CHEMISTRY    AXD    PHYSICS    OF    THE    CELL 

found  in  plants,  in  which  tlie  ])r()niinent  feature  is  tlie  limiting-  wall, 
forming-  a  cell  to  enclose  a  fluid  content.  In  most  instances  the 
"cell"  answers  better  to  the  definition,  "a  mass  of  protoplasm";  but 
usage  makes  language,  and  no  possible  confusion  can  arise  from  the 
prevailing  universal  use  of  the  original  term,  except,  perhaps,  that 
the  term  is  prone  to  carry  with  it  the  thought  of  the  w^alls  of  the 
cell  being  much  more  prominent  than  they  really  are.  This  is  not 
so  unfortunate  a  result,  perhaps,  for,  as  we  shall  see  later,  the  limiting 
surfaces  of  the  cell,  even  when  too  thin  to  be  readily  demonstrable, 
may  play  a  much  more  important  part  in  cell  chemistry  than  their 
appearance  indicates. 

The  morphological  division  of  the  cell  into  cell  wall,  cytoplasm, 
nucleus,  and  nucleolus  can  hardly  be  followed  out  chemically,  for  if 
we  surmount  to  some  extent  the  difficulties  in  the  way  of  studying  the 
different  portions  separately,  we  find  that  the  differences  between  them 
are  rather  quantitative  than  qualitative.  And,  furthermore,  however 
different  the  cells  of  one  organ  or  tissue  may  appear  from  those  of 
another  organ  or  tissue  under  the  microscope,  when  analyzed  by  the 
chemical  methods  at  present  at  our  disposal  we  find  the  differences 
very  slight  indeed.  Certain  substances  are  found  in  every  living  cell, 
and  in  quantities  usuall}^  not  greatly  dissimilar;  hence  they  are  as- 
sumed to  be  the  most  important  constituents  of  protoplasm,  and  are 
sometimes  called  the  primary  constituents  of  the  cell.  ]\Iany  other 
secondary  constituents  may  also  be  present,  some  of  which  are  so 
nearly  universal  that  we  are  not  sure  but  that  they  really  are  primary 
cell  components;  such  are  fat  and  glycogen.  Others  are  characteris- 
tics of  certain  cells,  such  as  melanin  and  keratin,  or  specific  products  of 
cell  metabolism,  such  as  mucin  and  the  specific  enzymes.  The  great 
histological  and  chemical  differences  existing  between  different  tis- 
sues depend  often  on  these  secondary  products,  as  in  fat  tissue  and 
squamous  epithelium;  or  upon  the  intercellular  substance,  as  with 
connective  tissue,  cartilage,  bone,  etc.,  which  may  be  looked  upon  as 
pi'odiK'ts  of  cell  activity. 

Protoplasm,  as  the  term  is  generally  used,  includes  the  various 
primary  constituents  with  the  fluids  permeating  or  dissolving  them, 
but  does  not  include  the  more  conspicuous  secondary  constituents, 
such  as  fat  droplets,  pigment  granules,  etc..  nor  the  cell  membrane 
when  such  exists.  Evidently  it  is  a  very  indefinite  term,  to  be  avoided 
as  much  as  possible.  ])articularly  because  of  the  confusion  as  to 
whether  it  includes  the  nucleus  or  not,  difrerent  authors  difVering  in 
this  i'es|)ect  in  their  usage  of  the  word. 

CHEMISTRY  OF  THE  ESSENTIAL  CELL  CONSTITUENTS 

To  enuinei'ate  the  primary  or  essential  constituents  of  tlu^  cell  al)- 
solntely    is   not    ])ossil)le,    for   the    rapid    advances    in    cheiiiisti'v    nuiy 


PROTEINS  19 

alter  all  classifications  without  warning,  but  practically  they  may  be 
grouped  under  the  headings  of  proteins,  lipins,  salts,  and  water,  and 
no  attempt  will  be  made  to  give  here  more  tiian  the  most  essential 
features  concerning  each. 

PROTEINS  2 

In  the  last  few  years  we  have  obtained  something  approaching  a 
scientific  understanding  of  the  chemical  nature  of  this  great  group  of 
the  most  highly  C()mi)lex  bodies  known  to  chemistry,  although  we 
are  still  not  in  a  position  where  it  can  be  positively  said  just  how  the 
various  components  of  the  molecule  are  united,  or  in  exactly  what 
proportion ;  and  we  are  still  farther",  perhaps,  from  the  point  of 
synthesizing  a  full-fledged  protein  molecule.  But  it  is  certain  that 
the  problems  regarding  the  underlying  principles  of  the  formation 
and  structure  of  the  giant  protein  molecule  are  nearing  solution. 
Our  information  has  been  obtained  almost  exclusively  through  studies 
of  the  products  obtained  by  splitting  up  the  proteins,  for  as  yet 
relatively  little  has  been  accomplished  through  synthesis.  Proteins 
can  be  decomposed  by  the  action  upon  them  of  acids  or  alkalies  in 
various  concentrations,  by  superheated  steam,  by  digestive  ferments, 
and  by  bacteria.  The  products  obtained  in  these  different  ways  are 
not  all  the  same,  for  some  substances  may  be  formed  by  oxidation,  re- 
duction, decomposition,  combination,  or  condensation  of  the  various 
products  of  simple  cleavage,  and  it  is  necessary  to  distinguish  between 
the  primary^  cleavage  products  (those  which  exist  as  radicals  within 
the  molecule)  and  the  secondary  products  (those  not  existing  pre- 
i'ormed  in  the  molecule  but  formed  by  transformation  of  tlie  primary 
products).  This  can  usually  be  done,  and  it  is  found  that  so  far  as 
the  primary  products  are  concerned,  it  makes  little  difference  which 
method  of  cleavage  (or  hydrolysis,  since  in  the  splitting,  water  is  com- 
bined with  the  organic  substances)  is  used. 

At  first  the  proteins  split  up  into  compounds  still  possessing  many 
of  the  features  of  the  typical  protein  molecule,  such  as  albumoses  and 
peptones,  and  these  bodies  are  then  further  resolved  into  simple  sub- 
stances, which  are  not  aggregates  of  several  smaller  molecules  as  are 
the  proteins,  and  which  can  be  obtained  in  pure  crystalline  forai.  No 
matter  which  method  is  used  we  find  the  process  going  through  these 
stages,  and,  as  before  mentioned,  the  primary  crystalline  products 
obtained  are  practically  the  same  quantitatively  as  well  as  qualita- 
tively. Some  methods,  e.  g.,  bacterial  decomposition,  however,  lead  in 
the  end  to  more  profound  or  different  decomposition  of  the  cleavage 
products  into  secondary  substances.     The  similarity  of  the  results  ob- 

2  For   the  complete    literature   of   this   subject   see   :Mann's    "Chemistry   of   the 
Proteins."  New  York.  1000;   "The  Chemical  Constitution  of  the  Proteins,"  Plim- 
mer,  London,  1012:   "The  General  Character  of  the  Proteins,"  Schryver,  London 
1912:  "Tlie  Vegetable  Proteins."  Osbnrne,  Tx)n(lon,  1010;    (all  the  last  tliree  bein<r 
in  the  series  of  "Monographs  on  .Biochemistry") . 


20  THE    CHEMISTRY    AND    PHYSICS    OF    THE    CELL 

tained  in  these  different  ways  indicates  that  there  are  definite  lines  of 
cleava<i'e  in  the  protein  molecnle  along  which  separation  takes  place, 
independent  of  the  nature  of  the  agency  at  work,  and  that  the  sub- 
stances obtained  represent,  as  the  Germans  figuratively  say,  the 
"building  stones"  of  the  entire  molecule. 

These  substances  all  have  in  common  one  important  point :  each  one 
is  an  acid,  which  has  a  NIIo  group  substituted  for  a  hydrogen  atom  on 
the  carhon  nearest  the  acid  radical  (the  a-position).  It  makes  no  dif- 
ference what  the  rest  of  the  radicals  are,  whether  they  are  simple 
chains  (leucine),  or  members  of  the  cyclic  or  aromatic  series  (ty- 
rosine), or  sulphur-containing  bodies  (cystine),  withou.t  exception 
this  relation  of  a  NIL  group  to  an  acid  radical  is  constant,  as  in  this 
formula:  XH, 

/ 
R— CH— COOH. 

Through  this  arrangement  every  one  of  the  constituents  of  the 
protein  molecule  is  provided  with  a  group  with  a  strong  basic  char- 
acter and  a  group  with  a  strong  acid  character,  and  hence  it  is  pos- 
sible for  them  to  unite  with  one  another  in  indefinite  numbers,  and, 
because  of  the  great  variety  of  individuals,  in  practically  an  infinite 
number  of  combinations.  It  is  believed  that  it  is  in  just  this  way  that 
the  protein  molecule  is  built  up.  By  artificially  uniting  various 
cleavage  products  Emil  Fischer  has  succeeded  in  producing  large 
molecules  made  up  of  several  amino-acid  radicals  (called  by  him 
"polypeptids")3  which  show  some  of  the  characteristics  of  the  pep- 
tones, and  this  is  the  nearest  that  investigators  have  yet  come  to  synthe- 
sizing a  protein  molecule.  The  union  is  accomplished  by  the  split- 
ting off  of  water,  corresponding  to  the  addition  of  water  that  occure 
when  the  protein  molecule  undergoes  cleavage.  It  may  be  illustrated 
by  showing  the  formation  of  the  simplest  polypeptid,  gbjcijighjcine. 

NH,  o  NH,       o 

CHo  — C— lOH   +   HI  HN  — CH;  — COOH  =  CH2  —  C  —  HN  —  CHo  —  COOH   +   H:0. 
(glycocoll)  (glycocoll)  (glycylglycine) 

For  these  reasons  it  is  believed  that  the  protein  molecule  consists 
of  great  nunibers  of  amino-acid  groups,  comhined  ivith  one  another 
through  their  hasic  and  acid  radicals,  and  that  the  various  proteins 
are  different  from  one  another  because  they  contain  different  num- 
bers or  varieties  of  amino-acids.  For  example,  the  glohin  of  hemo- 
globin yields  no  glycocoll  on  hydrolysis,  while  gelatin  yields  16.5  per 
cent.  On  the  other  hand,  gelatin  is  free  from  tyrosine.  Some  of 
the  protamins  (proteins  obtained  chiefly  from  spermatozoa)  yield  as 
high  as  58  to  84  per  cent,  of  arginine,  while  the  simpler  amino-acids 
Avith  but  one  N  (mono-amino-acids)  are  scanty,  and  most  varieties 
are  lacking. 

It  will  be   noticed  that  when   two   amino-acids  unite,    as   seen    in 

3  Reviewed  by  Fisolior,  in  Rer.  cliMit.  Cliom.  Gosoll.,  1906   (."^9).  ri.50. 


PROTEINS  21 

the  formation  of  glycylglycine,  an  acid  radical  and  a  basic  radical  are 
still  left  free.  In  this  may  be  seen  the  explanation  of  the  peculiar 
amphoteric  nature  of  proteins.  As  long  as  these  two  groups  are  free 
the  proteins  can  combine  with  either  acids  or  bases,  as  they  are  well 
known  to  do,  and  hence  they  react  as  either  acids  or  bases  under  dif- 
ferent conditions. 

It  must  not  be  imagined  that  the  structure  of  the  complete  mole- 
cule is  simply  a  long  straight  cliain  of  amino-acids  joined  only  in  the 
same  way  as  are  the  components  of  glj^cylglycine.  The  existence  of 
the  diamino-acids,  of  the  benzene  rings,  of  hydroxyl  groups,  (as  in 
serine  or  tyrosine),  of  ring  compounds,  (as  pyrrolidine  carboxylic 
acid),  of  substances  with  two  acid  groups,  (as  glutaminic  and  aspar- 
tic  acid),  adds  complications  to  the  formation  until  it  is  impossible 
to  estimate  just  liow  all  the  various  building  stones  may  be  arranged. 
"We  must  bear  in  mind  the  size  of  the  protein  molecule,  which  Hof- 
meister  has  estimated  (for  serum  albvimin)  as  having  a  molecular 
weight  of  10,166,  and  for  hemoglobin  the  molecular  weight  has  been 
estimated  at  16,669.^  Within  such  a  "giant  molecule"  there  is  room 
for  variety  almost  beyond  computation. 

The  Proteins  of  the  Cell. — By  physiological  chemists  proteins  are 
classified  into  simple  proteins,  of  which  egg  and  serum  albumin  are 
types;  and  compound  proteins,  which  are  characterized  by  having 
some  special  non-protein  group  which  can  be  split  off,  leaving  behind 
a  characteristic  protein  residue,  e.  g.,  nucleo-proteins,  glyco-proteins. 
As  primary  cell  constituents  the  following  varieties  of  proteins  may 
be  mentioned ;  albumin,  globulin,  nucleo-protein,  nucleo-albumin  or 
phospho-protein,  and  insoluble  proteins.  At  one  time  it  was  thought 
that  cytoplasm  consisted  chiefly  of  albumin,  like  white  of  egg,  but 
we  now  know  that  this  forms  but  a  small  part  of  the  cell  proteins, 
often  occurring  only  as  traces.  It  is  held  by  some  that  true  albumin 
occurs  only  as  a  building  or  intermediate  cleavage  product  of  the 
more  complicated  forms  of  cellular  proteins,  and  is  itself  of  relatively 
sliglit  importance  in  cell  life,  not  participating  in  chemical  changes 
except  as  a  food-stuff. 

Albumins  are  characterized  chiefly  by  their  greater  solubility  in 
water,  and  in  being  less  easily  precipitated  than  most  proteins.  They 
seem  to  be  a  fundamental  type  of  proteins.  The  three  forms  of  al- 
bumin that  have  been  described  in  animal  tissues  or  products  are  egg- 
albumin,  lactalbumin  of  milk,  and  serum  albumin ;  probably  cell  al- 
bumin is  most  closely  related  to  the  last,  and  what  has  been  described 
as  cell  albumin  is  perhaps  in  many  cases  but  serum  albumin  that  has 
been  imperfectly  removed. 

4  Robertson  (Jour.  Biol.  Chem.,  1909  (6),  105)  susrgests  that  these  high  molec- 
\ilar  weights  represent  polymerization,  the  single  protein  molecules  being  much 
smaller;  casein  salts  in  neutral  solutions  have  a  molecular  weight  of  but  2000, 
according  to  his  investigations. 


22  THE    CHEMISTRY    AXD    PHYi^ICS    OF    THE    CELL 

Globulins  also  occur  in  all  cells,  but  in  small  amounts  in  most  ani- 
mal cells  except  the  muscles,  whose  chief  proteins  belong  to  this  or  a 
closelj'  related  group.  The  globulins  are  quite  similar  to  the  albu- 
mins, so  that  there  is  really  no  sharp  line  between  the  two  groups. 
Their  insolubility  in  water  separates  them  from  albumins,  and  their 
solubility  in  dilute  neutral  salt  solutions  from  the  more  complex  pro- 
teins. An  important  feature  of  the  globulins  is  the  low  tempera- 
ture at  which  they  coagulate — some  so  low  that  Halliburton  ^  believes 
it  possible  that  they  may  be  coagulated  within  the  cells  during  high 
fevers. 

Hammarsten  has  long  maintained  that  simple  proteins  form  a 
relatively  insignificant  part  of  the  cytoplasm,  in  opposition  to  the 
once-prevalent  view  that  the  nucleo-proteins  were  limited  to  the 
micleus,  and  that  the  cytoplasm  was  chiefly  albumin  and  globulin. 
The  general  trend  of  opinion  as  influenced  by  the  results  of  researches 
has  been  favorable  to  his  contentions,  and  we  shall  probably  not  be  far 
wrong  in  accepting  his  statement  that — ''The  chief  mass  of  the  pro- 
tein substances  of  the  cells  does  not  consist  of  proteins  in  the  ordinary 
sense,  but  consists  of  more  complex  phosphorized  bodies,  and  that  the 
globulins  and  albumins  are  to  be  considered  as  nutritive  materials  for 
the  cells  or  as  destructive  products  in  the  chemical  transformation  of 
the  protoplasm. ' '  ^ 

Nucleo-proteins  are  probably  the  most  important  constituents  of  the 
cell,  both  in  quantity  and  in  relation  to  cell  activity.  In  structure 
the  nucleo-proteins  are  very  complex,  as  indicated  by  the  different 
products  yielded  on  hydrolytie  cleavage  of  the  molecule.  Further- 
more, there  are  many  varieties,  depending  both  upon  the  nature  and 
proportions  of  the  component  parts.  They  may  be  described  as  con- 
sisting of  two  primarj^  constituents — (1)  nucleic  acid  and  (2)  a  pro- 
tein body,  in  chemical  combination  with  each  other  like  a  salt.  The 
term  nucleic  acid  covers  a  large  group  of  substances,  which  are 
characterized,  on  the  one  hand,  by  their  frequent  occurrence  bound 
with  proteins,  and,  on  the  other  hand,  by  their  yielding  phosphoric 
acid  and  purine  bases,  pyrimidines  and  pentoses  or  hexoses  on  cleav- 
age. Diagrammatically  the  manner  of  cleavage  of  the  nucleo-proteins 
may  be  indicated  as  follows: 

Niicleo-protoin 

nuclcin  7         protein 

/\ 

nucleic  acid  ])rotcin 

/\ 

phosphoric  a<'i(l  ])minc  bases,   jiyrimidines   ajid   carltohydrates.  ^ 

5  Hallilmrton  and  ISfott,  Archives  of  Ncurolopy.  100.3  (2).  727;  also  see  TTalli- 
biirton's  "Chemistry  of  Muscle  and  Nerve." 

<i  See  Kossel,  Miincli.  nied.  Woch..  1011    (.')8),  fi.''). 

7  Proltahly  nncleiii  should  he  considered  as  merely  one  variety  of  niicleoin'otein, 
with  less  protein  llian  the  other  varieties. 


FATfi  AXD   IJI'O/ns    (/.//'/ V.ST)  23 

111  tlie  cell  the  imcleo-proteins  probably  exist  partly  as  solid  struc- 
tures, c.  g.,  the  chromatin  framework  of  the  nucleus,  and  partly  dis- 
solved in  the  plasma.  An  interesting  phenomenon  is  the  alteration  in 
the  chromatin  nueleo-proteins  durin<>:  cell  division,  when  they  seem  to 
lose  part  of  the  combined  protein  and  approach  more  nearly  pure 
nucleic  acid — just  as  inorganic  salts  occur  with  the  acids  and  bases 
saturating  each  other  more  or  less  incompletely,  e.  g.,  mono-,  di-,  and 
tribasie  ])li()sphates.  in  this  we  have  a  chemical  explanation  of  the 
intensity  of  the  staining  of  dividing  nuclei  by  basic  dyes.'' 

Phosphoproteins.  Of  these,  by  an  luifortunate  similarity  of  name, 
the  so-called  "nucleo-albumins"  are  often  confused  with  nueleo-pro- 
teins by  non-chemical  writers,  a  difficulty  increased  by  an  actual  re- 
semblance to  the  extent  that  they  also  yield  phosphoric  acid,  and  are 
somewhat  similar  in  solubility  and  digestibility.  They  are  essentially 
different,  however,  in  that  they  do  not  yield  nucleic  acid  or  purine 
bases  on  cleavage.  Probably  members  of  this  group  are  also  constant 
components  of  cells. 

Glycoproteins  (or  gluco-proteins)  and  pliospho-glycoproteins  are 
also  believed  to  occur  frequently  or  constantly  in  protoplasm.  They 
are  compounds  of  proteins  with  a  sugar  or  sugar-like  group,  which 
probably  usually  contains  nitrogen,  thus  differing  from  the  ordinaiy 
hexoses  and  pentoses. 

Insoluble  proteins,  or  bodies  resembling  the  coagulated  proteins  in 
their  lack  of  solubility  in  various  fluids,  are  left  behind  after  the 
other  proteins  have  been  extracted  from  the  cells.  Their  signitieance 
is  not  known :  whether  to  a  large  extent  artificially  produced  or 
whether  a  normal  structural  element  of  the  cell. 

FATS  AND  LIPOIDS    (LIPINS) 

Lipoids  is  a  term  in  common  use  but  of  indefinite  significance ;  most 
usually  it  comprehends  the  intracellular  substances  which  are  soluble 
in  ordinaiy  fat  solvents,  but  which  are  not  simple  fats  or  fatty  acids, 
lecithin  and  cholesterol  being  the  most  important  of  the  lipoids.  For 
the  entire  group  of  fats  and  lipoids  the  term  lipins  has  been  proposed 
by  Gies  and  Rosenbloom.  Lipoids  and  ordinary  fats,  that  is  lipins, 
occur  in  all  cells,  but  their  demonstration  is  not  always  readily  pos- 
sible. The  microscopic  appearance  of  a  cell,  even  when  special  stains 
for  fat  are  used,  gives  no  correct  idea  of  the  amount  of  lipins  actually 
present.  Thus  normal  kidneys  contain  15  to  18  per  cent,  in  their  dry 
substance,  but  none  of  this  can  be  detected  readily  with  the  micro- 
scope. A  kidney  which  seems  microscopically  the  site  of  marked  fatty 
degeneration  may  show  no  more  fat  when  examined  chemically  than  a 
normal  kidney,  which  in  section  appears  to  be  quite  free  from  fat. 
This  is  because  some  of  the  intracellular  fat  is  bound  chemically  with 

8  The  oheniistrv  of  the  niicleo-proteins  is  discussed  in  the  chapter  on  Frio  Acid 
Metabolism  and  Gout.  Chap.  xxi. 


24  77//;    CHEMISTRY    AND    PHYSICS    OF    THE    CELL 

the  proteins,)  and  when  so  bound  it  cannot  be  seen,  nor  can  it  be 
stained  by  the  dyes  ordinarily  used  for  that  purpose ;  onl}-  when  de- 
generative changes  of  certain  khids  have  liberated  it  from  combina- 
tion does  it  become  visible  and  stainable  by  ordinary  methods  (Rosen- 
feld).  By  the  special  fixation  method  of  Ciaccio  the  fatty  compounds 
of  even  normal  cells  may  be  made  stainable  (Bell),^  showing  that 
the  so-called  masked  fat  is  really  in  a  not  altogether  invisible  form. 
Whether  the  intracellular  fat  has  any  function  other  than  that  of 
serving  as  a  food-stuff  is  not  known,  but  there  can  be  no  question  of 
the  importance  of  the  phosphorized  fats,  or  phospholipins. 

Lecithin  is  a  primarj^  cell-constituent,  and  is  probably  important 
both  in  metabolism  and  physically.  liammarsten  regards  it  as  con- 
cerned in  the  building  up  of  the  nucleus.  As  will  be  shown  later, 
many  of  the  most  essential  physical  properties  of  the  living  cell  de- 
pend upon  the  presence  in  it  of  lipoids,  of  which  lecithin  is  appar- 
ently the  chief.  Of  the  ether-soluble  substances  in  the  heart,  for  ex- 
ample, 60  to  70  per  cent,  is  lecithin,  which  constitutes  about  8  per 
cent,  of  the  dry  weight  of  the  myocardium, 

There  are  several  varieties  of  lecithin,  depending  upon  the  fatty 
acid  radical  they  contain.  The  structural  formula  of  one  lecithin, 
stearyloleyl  lecithin,  is  as  follows: 

CH,— 0— C— H3,0 

CH— 0— Cis— H33O 

I 
CH„— 0— PO— OH 

I 

0— CH„— CH2— N  =E    (CHala. 

OH 

It  differs  from  ordinary  fats,  therefore,  in  having  two  special  groups, 
one  the  phosphoric  acid,  the  other  the  choline  radical,  which  last  may 
be  of  some  importance  in  pathological  processes.  In  its  ]ihysical 
properties  it  is  quite  similar  to  tlie  ordinary  fats,  although  it  forms 
even  finer  emulsions  in  water,  which  are  .practically  colloidal  solutions 
(W.  Koch). 

Cephalin,  a  closely  related  body  differing  in  having  but  one  methyl 
grouj),  is  also  pi-obably  as  M^dely  spread  in  the  tissues  as  lecithin, 
according  to  Koch  and  Woods. ^° 

Cholesterol,  which  is  another  lipoiil,  is  nearly  as  universally  present 
as  lecithin. ^^  It  exists  both  free  and  in  combination  with  fatty 
acids,  for  chf)lesterol  is  an  alcohol  and  not  at  all  similar  to  the  fats 
chemically,  aitliough  very  similar  physically.  The  empirical  formula 
is   CoJT.,,011   or   C,-H.t,blT,   and   it   is   ivJal.'d    I0   llic    terpenes.     It 

n.Tour.  ^^\od.  Eos.,  mil    (10),  539. 

if>.Tour.  Biol.  C'hoin.,  100.5    (1),  20.1. 

11  Recent  literature  piven  liv  Clikin,  llioclicm.  Cciilr..  lOOS    (7).  2Sn. 


CARBOHYDRATES  25 

seems  to  be  relatively  inert  chemically,  and  therefore  is  probably  im- 
portant only  because  of  its  effect  on  the  physical  properties  of  the 
cells.  By  some  it  is  considered  to  be  a  decomposition  or  cleavage 
product  of  the  proteins,  which  is  in  accordance  witli  its  abundance 
in  masses  of  old  necrotic  tissue,  e.  <j.,  atheromatous  masses,  old  in- 
farcts, and  old  exudates. 

Protagon,  which  name  probably  covers  a  <;Toup  of  nitrogenous, 
phosphorized  bodies,  (Gies),^-  occurs  in  many  or  all  cells,  but  espe- 
cially in  the  nervous  tissues.  The  properties  of  protagon  are  in  gen- 
eral similar  to  those  of  the  other  lipoids,  but  its  exact  composition  is 
too  uncertain  to  permit  of  surmises  as  to  its  special  purpose. 

Doubly  Refractive  Lipoids  and  Myelins.^' — In  practically  all  nor- 
mal tissues  there  are  present  droplets  of  lipoid  nature  which  are 
characterized  b}^  showing  prominent  crosses  when  examined  with 
crossed  Nicol  prisms  (anisotropic),  the  adrenal  and  corpus  luteum 
containing  them  most  abundautl}^.  Chemically  they  seem  to  be  mix- 
tures of  various  lipoids  in  inconstant  proportions,  but  probably  the 
anisotropic  character  is  most  usually  dependent  upon  the  presence  of 
cholesterol  esters.  The  term  myelin  was  first  applied  by  Virchow 
to  peculiar  fatty  substances  found  in  various  normal  and  pathological 
tissues,  because  they  showed  physical  characters  similar  to  those  of  the 
myelin  substance  of  nerves,  but  as  many  of  these  substances  are  doubly 
refractive,  or  can  be  easily  made  so,  some  authors  use  the  term  myelin 
as  if  it  were  synonymous  with  doubly  refractive  lipoids.  There  are, 
however,  myelins  which  are  not  always  doubly  refractive,  and  also 
double  refractive  lipoids  which  do  not  swell  up  in  water  to  form 
myelin  figures,  etc.,  as  is  characteristic  of  true  myelins.  Chemically, 
however,  the  myelins  and  doubly  refractive  substances  are  probably 
related,  consisting  of  mixtures  of  cholesterol,  cholesterol  esters,  lecithin 
and  perhaps  soaps,  in  varjdng  proportion.  They  will  be  considered 
further  in  discussing  Fatty  ^Metamorphosis,  Chap,  xiv, 

CARBOHYDRATES 

The  third  great  class  of  food-stuffs,  the  carbohydrates,  is  represented 
in  the  cell  by  pentoses  and  Jiexoses  combined  with  proteins  and  with 
lipoids,  and  also  by  glycogen,  which  exists  free.  Glycogen  is  a  rather 
difficult  substance  to  isolate  in  minute  cpiantities  and,  therefore,  al- 
though it  is  not  found  in  all  cells  by  our  present  methods,  yet  it  may 
well  be  that  it  is  a  constant  constituent  of  the  protoplasm.  There 
is  no  evidence,  however,  that  it  is  anything  more  than  a  source  of 
heat  and  energy  to  the  cell.  Its  properties  and  occurrence  will  be 
considered   more   fully   in   the   discussion    of   glycogenic   infiltration. 

12  Jour.  Biol.  Chem.,  1905    (1),  50. 

13  See  Adami,  Jour.  Amer.  Med.  Assoc,  1007  (48),  463:  Karwicka.  Ziepler's 
Beitr.,  1911  (50),  437;  Schultze,  Ergebnisse  Pathol.,  1909  (13,  pt.  2),  253;  Bang, 
Ergebnisse  Phvsiol.,  1907    (6),  131;   1909    (8),  463. 


26  Tin:    CHEMISTRY    AM>    I'lIYHICH    OF    THE    CELL 

Since  glj^eogen  is  formed  from  dextrose  and  is  constantly  breaking 
down  into  dextrose,  it  is  probable  that  the  latter  is  also  constantly 
present  in  the  cells. 

INORGANIC   SUBSTANCES 

Up  to  this  point  the  substances  of  the  cytoplasm  that  have  been  dis- 
cussed have  all  been  organic  compounds  which  do  not  naturally  exist 
independent  from  living  or  once  living  cells,  yet  the  inorganic  sub- 
stances of  the  protoplasm  are  also  of  vital  importance.  As  Mann 
says,  "so-called  pure  ash-free  proteins  are  chemically  inert,  and,  in 
the  true  sense  of  the  word,  dead  bodies.  AVhat  puts  life  into  them 
is  the  presence  of  electrolytes."  The  various  salts  of  i)otassium,  so- 
dium, calcium,  magnesium,  and  iron  which  all  cells  contain  do  not  exist 
merely  dissolved  in  the  water  of  the  cell,  but  in  part  they  are  com- 
bined with  tlie  organic  constituents  of  the  protoplasm.  They  are  not 
combined  as  simple  additions  of  the  salts  to  the  proteins ;  but  ions,  both 
anions  and  cations,  are  united  in  chemical  combination  to  the  large  pro- 
tein molecule  (ion-proteins).  Possibly  the  proteins  participate  in  vi- 
tal chemical  processes  only  as  ion  compounds  with  inorganic  elements. 
It  is  extremely  difficult,  indeed  almost  impossible,  to  secure  proteins 
entirelj^  free  from  inorganic  substances  (ash-free  proteins).  The  fact 
that  inorganic  substances  are  held  in  the  cells  chemically  rather  than 
by  simple  diffusion  into  them  from  the  surrounding  fluids  is  sho^^m 
by  the  great  difference  in  the  proportions  of  various  salts  in  the  cells 
and  in  the  extra-cellular  fluids.  Thus  potassium  is  nearly  always 
much  more  abundant  in  the  cells  than  in  the  tissue  fluids,  while  so- 
dium is  more  abundant  in  the  fluids.  Phosphoric  acid  is  also  more 
abundant  in  the  cells,  and  chlorin  in  the  plasma.  In  cells  iron  seems 
to  exist  chiefly  in  combination  with  the  nucleo-proteins.  These  mat- 
ters will  be  taken  up  in  greater  detail  in  considering  the  physical 
chemistry  of  the  cell." 

THE  PHYSICAL   CHEMISTRY  OF  THE  CELL  AND  ITS   CONSTIT- 
UENTS "^ 

From  the  standpoint  of  physical  chemistry  the  cell  consists  of 
a  collection  of  colloids  and  crystalloids,  electrolytes  and  non-electro- 
lytes, dissolved  in  water,  in  lipoids,  and  in  each  other,  surrounded  by 
a  semipermeable  membrane,  and  perhaps  subdivided  by  similar  mem- 
branes or  surfaces.  Physical  chemical  processes,  as  we  shall  see  later, 
play  an  all-important  part  in  the  life  phenomena  of  the  cell,  and 
therefore  some  s])ace  may  l)e  occupied  ])rofitably  in  explaining  tlie 
nature  of  these  changes  and  of  the  substances  that  particii)ate  in  them. 

14  Soe  ^raf'.alluni  on  ^tici'oclioinistry,  l^rfxobnissc  Pliysiol.,   lOOS    (7),  5;">2. 
i-t'i  S(>e   I'.ayliss,  ■"I'liiiciph's  of  (icMicral    Pliysiok)gy,"'  London,    1015,  for  a  more 
extensive  diseiissinii  of  lliese  (opii-s. 


vinsTM.t.ofDs  A\n  Tin:/!,-  /'i!ori:irrfi:s  27 

CRYSTALLOIDS  AND  THEIR  PROPERTIES 

Crystalloids,  or  substances  tliat  tend  under  favorable  conditions  to 
form  crystals,  aiul  whicli  diffuse  readily  tlu'()u<;h  most  diffusion  mem- 
branes, form  a  I'elatively  small  part  of  tlie  total  mass  of  tbe  cell,  but 
they  are  fully  as  essential  as  the  colloids.  The  chief  representatives 
of  this  group  that  are  found  usually  or  constantly  in  the  cell  are  the 
inorganic  salts,  sugar,  and  the  innumerable  decomposition  products 
of  the  proteins,  including  particularly  urea,  creatine,  purine  bases, 
amino-acids,  etc.  ^Most  of  these  are  by  no  means  so  characteristic  of 
living  things  as  are  the  colloids,  sometimes  occurring  quite  inde- 
pendently of  a  cellular  origin,  which  the  proteins  never  do.  The  inor- 
ganic salts  in  particular  seem  quite  foreign  to  living  processes,  and 
as  they  enter  and  leave  the  body  practically  unchanged  they  are 
evidently  not  a  source  of  energy  through  chemical  change.  Their 
importance  to  the  cell  lies  almost  entirely  in  their  physical  or  physico- 
chemical  properties.  The  organic  crystalloids,  although  of  initritional 
value,  also  have  physical  properties  in  some  respects  similar  to  those 
of  the  inorganic  crystalloids,  and  therefore  to  this  extent  they  exert 
similar  influences,  but  the  essential  difference  between  the  organic 
and  the  inorganic  crystalloids  is  that  all  the  latter  are  electrolytes, 
while  many  of  the  organic  crystalloids  that  occur  in  cells  are  non- 
electrolytes.  The  importance  of  this  distinction  lies  not  in  the  utility 
or  non-utility  of  these  substances  as  conductors  of  electrical  cur- 
rents in  the  ordinaiy  sense,  but  rather  on  the  existence  of  those 
properties  which  determine  their  conductive  ability.  Electrical  con- 
ductivity is  an  index  of  ionization,  and  upon  ionization  depends  the 
chief  influence  of  the  electrohi:es  upon  vital  activities.  The  impor- 
tance of  this  process  of  dissociation  or  ionization  lies  in  the  fact  that 
with  most  substances  no  chemical  reaction  can  occur  while  the  sub- 
stance is  in  the  non-ionized  state.  The  chemical  properties  of  ioniza- 
ble  substances  are  produced  largely  by  the  ions  they  liberate  on 
dissociation.  As  a  consecjuence,  the  physiological  effects  of  electro- 
lytes are  due  to  their  ionic  condition,  and  through  the  ions  that  are 
present  in  the  cell  many  of  its  various  chemical  processes  are  brought 
about.  Not  all  substances  ionize  with  the  same  readiness,  which 
causes  a  great  difference  in  their  properties.  The  reason  that  acetic 
acid  is  a  weaker  acid  than  hydrochloric  acid  is  that  it  does  not 
ionize  to  such  an  extent,  and  so  a  corresponding  quantity  does  not 
introduce  as  large  a  number  of  hydrogen  ions  into  a  solution. 
Larger  molecules,  as  a  rule,  ionize  less  than  smaller  ones  of  similar 
nature,  e.  g.,  stearic  acid  ionizes  less  than  acetic  acid  and  therefore 
is  a  weaker  acid.  Likewise  the  properties  of  a  substance  which 
depend  upon  its  ions  will  be  less  marked  when  it  is  in  a  solvent  that 
produces  little  ionization.  For  example,  bichloride  of  mercury  owes 
its  antiseptic  properties  to  the  Hg  ions  that  it  sets  free  when  in  solu- 


28  77//;  ciiKMisTny    wn  riivs/cs  of  the  cell 

tion.  It  is  woll  known  tliat  solutions  of  mercury,  and  for  tliat  matter 
most  other  antisp]:)tics,  are  much  less  actively  germicidal  in  alcohol 
than  when  in  water,  because  their  ionization  is  less  in  alcohol;  and  the 
germicidal  properties  decrease  as  the  proportion  of  alcohol  increases, 
until  the  germicidal  effect  of  the  mixture  is  no  greater  than  that  of 
alcohol  alone  in  the  same  strength. 

If  we  had  no  electrolytes  in  the  cell,  electric  charges  could  not  be 
carried  about  in  it,  and  hence  chemical  reactions  could  not  occur.  It 
is  this  fact  that  makes  the  inorganic  salts  of  such  vital  importance 
to  the  cell  life.  To  repeat  Mann's  words,  it  is  the  electrolytes  that  put 
life  into  the  proteins.  AYater  itself  is  almost  absolutely  non-dissoci- 
ated, and  proteins  so  little  that  for  some  time  it  w^as  doubted  if  they 
I'eally  did  ionize.  Probably  all  soluble  substances  do  dissociate  to  a 
certain  minimal  degree,  but  it  is  so  slight  for  most  of  the  constitu- 
ents of  the  cell  except  the  inorganic  salts  (the  organic  acids  and  alka- 
lies, and  a  few  dissociable  organic  products  of  protein  metabolism, 
occur  in  such  insignificant  amounts  as  to  be  almost  negligible)  that 
without  them  there  would  be  little  chemical  activity  possible,  and 
hence  life  would  be  absent  or  at  a  very  low  ebb  indeed.  As  before 
mentioned,  the  inorganic  salts  probably  exist  in  the  cell  not  only  as 
salts,  but  also,  and  perhaps  chiefly,  as  ions  and  ionic  compounds  w^ith 
the  cell  proteins.  For  the  most  part  it  seems  to  be  the  cations  that 
play  the  chief  role  in  forming  ion-protein  compounds,  althoiigh  un- 
doubtedly the  anions  do  combine  wdth  the  proteins  also,  and  in  some 
instances  they  exert  very  characteristic  and  important  effects;  e.  g., 
the  differences  between  the  effects  of  chlorides,  bromides,  and  iodides, 
or  of  CNH  as  compared  with  HCl,  both  of  wdiich  liberate  the  same 
cation  and  differ  only  in  their  anions. 

IMany  applications  of  the  facts  and  theories  of  ionization  have  been 
made  in  physiology,  and  a  few  applications  have  also  been  made  in 
pathology,  especially  the  relation  of  ions  to  edema,  to  diuresis  and 
glycosuria,  and  also  to  problems  of  immunity.  No  attempt  will  be 
made  here  to  go  further  in  the  observations  and  theories  concerning 
ionization  or  its  role  in  physiology,  but  for  more  extensive  informa- 
tion as  well  as  for  the  complete  bibliography  the  works  mentioned 
below  may  be  referred  to.^^  The  applications  in  pathology  will  be 
brought  out  as  the  subject  under  discussion  in  subsequent  chapters 
necessitates,  and  it  is  largely  to  facilitate  the  understanding  of  such 
references  that  this  brief  summary  of  the  subject  of  ionization  has 

15  "Physical  Chemistry  in  tlio  Service  of  Medicine."  Wolfsanir  Paul!,  transla- 
tion by  M.  TT.  Fischer,  New  York.  IflO".  "Physikalisohe  Chemie  der  Zelle  und 
der  Gewebe."  ITiiber,  Leipzig,  191;").  "Osniotische  Driick  iind  Tonenlehre  in  den 
medicinischen  Wissensehaften,"  Hamburjrer,  Wiesbaden.  "Studies  in  General 
Physiolopy."  Loeb,  University  of  Chicairo  Press,  1005.  "Dynamics  of  Livinjj 
]\ratter,"  Loeb,  Columbia  University  Press.  New  York,  1000.  Bottaz/i,  Er^eb- 
nisse  d.  Pliysiol.,  1008  (7),  IGl.  Spiro  and  J.  Loeb,  Oppenheimer's  "Handbuch 
der  Biochemie,"  1008   (2),  1-141. 


DIFFUSION  AM)  OSMOSIS  29 

been  introduced.  In  the  same  spirit  we  take  up  tlie  subjects  of  dif- 
fusion and  osmosis. 

Diffusion  and  Osmosis. — Although  the  non-electrolytes  do  not  ion- 
ize to  any  considerable  extent,  and  therefore  are  relatively  inactive 
chemically,  the  crystalloidal  non-electrolytes,  of  which  sugar  and  urea 
are  the  two  chief  examples  among  the  cell  constituents,  possess  in 
common  with  the  electrolytes  the  important  property  of  diffusion. 
By  this  process  the  exchange  of  chemical  substances  between  the  blood 
and  the  cell  is  brought  about,  by  it  the  chemical  composition  of  the 
different  parts  of  the  cell  and  between  different  cells  is  equalized, 
and  without  it  chemical  change  would  be  practically  impossible.  Dif- 
fusion occurs  most  simply  between  two  solutions  of  mil  ike  nature, 
or  between  a  solution  of  a  substance  and  the  solvent  alone,  when 
placed  directly  in  contact  with  one  another.  If  we  place  in  tlie  bot- 
tom of  a  cylindrical  vessel  a  solution  of  copper  sulphate  and  above 
it  some  water,  carefully  avoiding  mixing,  it  will  be  found  after  some 
time  tliat  the  fluid  has  become  equally  blue  throughout.  This  is 
brought  about  by  the  movement  of  the  dissolved  particles  which 
gradualh'  carries  them  through  the  entire  mass  of  fluid,  and  as  their 
migration  is  against  the  force  of  gravity,  they  evidenth'  accomplish 
work.  This  process  is  not  dependent  upon  ionization,  for  a  solution 
of  cane-sugar  or  of  urea  will  show  the  same  diffusion.  A  solution  of 
protein  or  other  colloid  does  so  much  more  slowly,  however,  indeed 
quite  imperceptibly. 

If  we  were  to  introduce  a  piece  of  filter-paper  between  the  water 
and  the  copper  sulphate  solution,  the  diffusion  would  go  on  the  same, 
the  pores  of  the  paper  permitting  the  passage  of  the  molecules  with- 
out hindrance.  If,  instead  of  filter-paper,  there  were  introduced  a 
sheet  of  some  substance  free  from  pores,  the  diffusion  would  be  much 
more  affected.  If  the  septum  was  of  such  a  nature  that  the  sub- 
stances in  solution  were  insoluble  in  it  (e.  g.,  glass),  diffusion  would 
of  necessity  stop ;  but  if  it  were  something  in  which  the  solvent  or  the 
solute  was  soluble,  such  as  a  gelatin  plate,  then  these  substances  would 
dissolve  in  it,  and  diffusing  through  its  substance  escape  into  the 
fluid  on  the  other  side.  The  last  example  indicates  the  conditions 
afforded  in  the  animal  cell,  and  also  in  the  usual  laboratory  diffusion 
experiments  when  the  membrane  is  generally  either  an  animal  mem- 
brane or  a  parchment  paper,  both  of  which  are  composed  of  colloids. 
Crystalloids  are  generally  soluble  in  colloids  and  hence  pass  through 
such  diffusion  membranes;  colloids  dissolve  but  slightly  in  colloids, 
and  hence  they  do  not  pass  through  a  diffusion  membrane  readily, 
and  are,  therefore,  but  very  slightly  diffusible. 

The  process  of  diffusion,  if  uninterrujited,  always  continues  until 
the  solution  is  of  exactly  the  same  composition  throughout.  If  on  one 
side  of  the  diffusion  membrane  there  is  a  substance  that  passes  through 
the  membrane   rapidly,   and   on   the   other  a  substance  that  passes 


30  Tin:  (iiEMisruY  wn  rinsicH  of  the  cell 

through  slowly  or  not  at  all,  tliere  will  soon  be  an  unequal  condition  on 
the  two  sides  of  the  membrane,  for  tlie  diffusible  substance  would  ac- 
cumulate in  equal  amounts  on  each  side,  while  the  non-diffusible  would 
remain  where  it  was.  On  one  side  there  would  then  be  more  material 
exerting  osmotic  pressure  than  on  the  other,  and  if  the  membrane 
were  flexible,  it  would  bulge  toward  the  opposite  side.  The  pressure 
is  supposed  to  be  due  to  the  bombardment  of  the  containing  walls  by 
molecules  or  ions  of  the  substances  in  solution,  and  hence  the  more 
molecules  and  ions  in  solution,  the  more  pressure.  When  equal  num- 
bers of  particles  are  on  each  side  of  the  partition,  the  pressure  is 
equalized.  It  is  quite  possible  to  have  membranes  readily  permeable 
to  one  substance  and  almost  entirely  impermeable  to  another;  such 
membranes  are  called  semipermeable.  To  produce  osmotic  pressure 
it  is  not  necessary  that  the  membrane  be  absokitely  impermeable  to 
any  of  the  substances — it  may  only  be  relatively  less  permeable  for 
the  solute  than  for  the  solvent.  If,  for  example,  we  fill  a  parchment 
bag  with  concentrated  sugar  solution,  tie  up  the  top  tightly  and  throw 
into  water,  it  will  swell  up  rapidly  and  eventually  burst.  But  if  the 
parchment  is  in  the  form  of  a  tube,  open  at  the  top,  and  the  lower 
end  is  placed  in  water,  the  amount  of  fluid  inside  the  tube  will  in- 
crease at  first,  but  eventually  the  sugar  will  diffuse  out  to  such  an 
extent  that  the  solution  is  of  the  same  concentration  inside  and  out- 
side of  the  tube,  and  the  column  of  fluid  will  again  become  of  equal 
height  on  both  sides.  These  results  indicate  that  the  water  passes 
through  the  membrane  more  rapidly  than  does  the  sugar,  but  that 
eventually  the  sugar  can  all  pass  through. 

Exacth'  similar  conditions  exist  in  cells,  particularly  plant  cells. 
The  typical  cell  of  plant  tissue  consists  of  a  distinct  wall,  usually 
cellulose  or  ehitin,  lined  internally  by  a  layer  of  protoplasm  which 
incloses  a  mass  of  aqueous  solution,  the  cell  sap,  containing  sugar  and 
various  other  solutes.  The  outer  wall  is  readily  permeable  by  water 
and  by  most  solutes,  whereas  the  protoplasmic  layer  inside  it  behaves 
like  a  semipermeable  membrane,  which  permits  water  to  pass  through 
readily  but  hinders  greatly  the  passage  of  most  solutes;  that  it  is 
somewhat  permeable  is  attested  by  the  fact  that  the  cell  sap  contains 
solutes  derived  fnmi  the  external  fluids.  As  a  result  of  this  arrange- 
ment there  is  a  constant  tendency  for  the  cavity  of  the  cell  to  be 
distended  by  water  and  for  the  solutes  within  it  to  exert  their  con- 
siderable pressure  upon  tlie  cell  wall.  Because  of  the  strengtli  of  the 
cellulose  layer  the  cell  can  withstand  great  pressures  that  would 
tear  apart  tlie  tender  protopla.smic  layer  that  really  determines  the 
osmotic  conditions,  just  as  in  the  exjierimental  membi-ane  the  clay 
cylinder  supports  the  delicate  i)recipitation  membrane.  It  is  the 
osmotic  pressure  that  causes  the  rigidity  oi-  turgor  of  plant  cells, 
and  explains  the  ability  of  a  tender  gi-ecii  shoot  to  hold  itself  up- 
j'iglit  oi-  horizontal  in  the  air;  and  it  is  the  foi-ce  that  enables  growing 


OSMOTIC  iMji:ssri;K  31 

roots  to  lift  <>reat  stoiu's  or  tear  apart  rocks  in  whose  clefts  they 
grow.  If  certain  plant  cells  are  placed  in  distilled  water,  the  pressure 
may  rise  to  such  an  extent  that  the  cells  burst,  and  it  was  throuf^h 
studyin<>:  this  ])lu'n()nieiion  that  Pfett'er  worked  out  the  basis  of  our 
present  knowledge  of  osmotic  pressure.  If  the  cell  is  placed  in  a  so- 
lution of  greater  concentration  than  its  cell  sap,  the  pressure  outside 
will  be  greater  than  that  inside  and  the  protoplasmic  membrane  will 
be  forced  away  from  the  cellulose  wall,  while  its  central  cavity  shrinks 
and  perhaps  disappears  entirely,  the  protoplasm  forming  a  ball  in  the 
center.  This  is  practically  what  occurs  when  a  plant  stem  is  cut 
and  it  "wilts" — the  water  is  removed  by  evaporation,  the  osmotic 
pressure  outside  the  cells  becomes  greater  than  that  inside,  and  the 
water  passes  out.  Likewise  when  a  plant  cell  dies  the  turgor  is  lost 
because  the  membrane  becomes  permeal)U\  and  so  pressure  soon  be- 
comes the  same  on  both  sides  of  the  cell  wall. 

I21  animal  cells  the  wall  is  not  so  highly  developed  as  in  plants, 
nor  is  it  backed  up  by  a  rigid  material  like  cellulose ;  indeed,  for 
many  animal  cells  there  is  no  well-defined  wall  and  the  protoplasm 
appears  to  be  naked.  Nevertheless  the  behavior  of  the  animal  cells 
indicates  that  they  do  possess  what  resembles  a  cell  wall,  in  that 
they  behave  when  in  solutions  as  if  the^^  were  surrounded  by  a  dif- 
fusion membrane.  The  degree  to  which  phenomena  of  this  nature 
are  shown  varies  with  different  cells;  with  red  corpuscles,  for  example, 
the  osmotic  pressure  influences  are  very  marked,  as  shown  by  the 
wrinkling  or  crenation  of  the  corpuscles  when  they  are  placed  in 
fluids  of  higher  concentration  than  the  blood  plasma,  and  by  their 
swelling  and  disintegration  with  escape  of  the  hemogiobin  (hemoly- 
sis) when  they  are  put  into  distilled  water  or  solutions  of  less  con- 
centration than  the  plasma.  Other  tissue  cells  seem  to  undergo  more 
or  less  alteration  from  changes  in  the  osmotic  pressure  in  the  fluids 
surrounding  them.  The  diffusion  membrane  that  surrounds  the  cell 
is  generally  not  well  defined,  and  for  most  cells  seems  to  be  but  a 
surface  condensation  of  the  protoplasm,  perhaps  formed  through  the 
effects  of  surface  tension.  It  seems  probable  that  this  surface  dif- 
fusion membrane  contains  a  large  proportion  of  cell  lipoids,  i.  e., 
cholesterol  and  lecithin  (for  the  red  corpuscles  this  is  practically 
certain)  ;  hence  substances  soluble  in  lipoids  penetrate  the  cell  read- 
ily, while  to  many  substances  insoluble  in  lipoids  the  cell  is  nearly 
or  quite  impermeable  (Overton).  Probably  the  wall  of  the  aninuil 
cell  is  not  so  nearly  semipermeable  as  is  that  of  the  plant  cell,  for 
nowhere  in  the  animal  body  do  we  get  such  turgor  in  the  cells  as 
we  see  in  plant  tissues.  Lacking  a  cellulose  wall,  animal  cells  could 
not  develop  such  an  internal  pressure  without  rupturing,  and  such 
a  process  of  rupturing  {plasinorrhexis,  plasmoptysis)  does  not  seem 
to  be  a  normal  occurrence  in  animal  tissues.  AVe  shall  be  most 
nearly  correct,  probably,  if  we  look  upon  the  animal  cell  as  possess- 


32  THE    CHEMIHTRY    AM)    I'llY^WS    OF    THE    CELL 

ing  a  delicate  diffusion  membrane  at  its  surface,  through  which  water 
passes  more  readily  than  do  most  crystalloids,  and  through  which 
colloids  pass  almost  not  at  all,  but  the  exclusion  of  each  of  these 
t^'pes  of  substances  is  merely  relative  and  not  absolute.  AVithin  the 
cell,  also,  the  colloids  probably  exist  as  a  more  or  less  well-developed 
emulsion,  so  that  we  have  here  a  practically  limitless  amount  of 
surface  formation  all  through  the  protoplasm;  such  a  structure  could 
permit  the  endless  number  of  reactions  of  a  living  cell  to  go  on 
side  by  side  in  the  same  cell.  Recent  studies  of  G.  L.  Kite  seem 
to  show  that  all  the  protoplasm  has  much  the  same  relation  to  solu- 
tions as  does  the  external  layer  or  cell  membrane,  for  he  found 
that  if  drops  of  solutions  which  can  penetrate  a  cell  from  outside 
be  injected  directly  into  a  cell  they  diffuse  through  it,  but  sub- 
stances which  camiot  penetrate  from  outside  are  also  unable  to 
diffuse  through  the  cell  after  they  have  been  injected  into  it. 

Since  osmotic  pressure,  exactly  like  gas  pressure,  is  presumably 
produced  by  the  bombarding  of  the  walls  of  the  container  by  parti- 
cles in  the  solution,  the  amount  of  pressure  will  vary  in  proportion  to 
the  number  of  particles  present.  With  such  substances  as  sugar 
and  urea,  the  non-electrolytes,  the  moving  particles  seem  to  be  mole- 
cules, and  so  a  solution  of  sugar  or  urea  will  produce  an  osmotic 
pressure  directly  proportional  to  the  number  of  molecules  it  con- 
tains. In  the  case  of  the  electrolytes,  however,  the  ions  produce 
pressure  as  well  as  the  molecules,  and  hence  an  electrolyte  in  solution 
will  produce  a  relativelj^  high  osmotic  pressure  as  compared  with  an 
equivalent  solution  of  a  non-electrolyte,  since  each  molecule  yields 
two  or  more  ions.  Colloids,  however,  exert  so  slight  an  osmotic 
pressure  that  it  is  difficult  of  detection;  this  probably  depends  on 
the  great  size  and  slight  motility  of  their  molecules.  In  the  many 
and  important  osmotic  processes  of  the  animal  organism,  therefore, 
the  colloids  take  no  part  except  in  helping  to  form  the  diffusion 
membrane,  and  in  preventing  the  diffusion  of  one  another.^®  It  is 
interesting  to  consider  also  that  colloids  under  ordinary  conditions 
do  not  greatly  modify  the  diffusion  of  crystalloids  through  a  solution 
containing  both  classes  of  matter.  The  fact  that  a  cell  is  full  of  dis- 
solved colloids  does  not  seriously  affect  the  osmotic  properties  of 
the  intracellular  crystalloids,  provided  the  colloids  are  not  condensed 
in  such  a  way  as  to  form  diffusion  nuMiibranes.  But  as  all  the  eleav- 
age  products  of  proteins  after  tliey  liave  passed  the  peptone  stage  are 
crystalloids,  by  decomposition  of  the  intracelhdar  proteins  the  os- 
motic pressure  may  be  greatly  raised.  As  long  as  the  cell  is  living 
there  can  be  no  constancy  in  composition,  for  metabolic  processes, 

10  Undor  cxpciiinontal  conditions  it  is  found  that  tlie  nature  of  the  membrane 
preatly  modifies  tlic  osmotic  ])reasure;  for  if  a  jjfiven  colloid  is  soluble  in  a  cer- 
tain membrane  and  a  certain  crystalloid  is  not,  the  colloid  will  ditVuse  tln-oujrh 
tlie  membrane  wliile  the  crystalloid  is  held  back.  (Kahlenberff,  Jour.  Physical 
Chem.,  1 !)()()   (10),  141.) 


OFMOTIC  I'fx'ESSrUE  33 

])}'  ])rodnc'ino-  from  proteins  that  liave  no  osmotic  pressure  erystal- 
!t)i(lal  substances  tluit  do  luive  osmotie  pressure,  cause  intracellular 
osmotic  conditions  to  be  continually  varjdng.  As  a  result,  streams 
of  diffusin<>-  particles  arc  m()vin<«-  a])out  in  every  direction,  settinjr  up 
new  chemical  reactions  and  consequent  new  osmotic  currents.  The 
{2:reater  the  ditlt'erence  in  osmotic  pressure  between  a  cell  and  its 
environs,  and  between  the  ditferent  parts  of  the  same  cell,  the  more 
powerful  the  osmotic  efiPects,  and  as  a  result  the  g^reater  the  capacity 
for  accomplish inji'  work.  The  storing-  up  of  insoluble  and  inditfusiblc 
forms  of  substance,  such  as  glycogen,  fat,  and  proteins,  is  an  im- 
portant factor  in  maintaining  inequalities  in  osmotic  pressure,  and 
in  this  way  of  increasing  work  Capacity. 

Indeed,  we  may  look  upon  cell  life  as  a  constant  attempt  at  the 
esta])lishment  of  equilibrium,  both  chemical  and  osmotic,  which  is 
never  achieved  because  the  move  towards  one  sort  of  equilibrium  is 
always  against  the  other.  All  the  food-stut?s — fats,  carbohydrates 
and  proteins— are  characterized  by  being  colloids  when  intact  and 
crystalloids  when  disintegrated,  thus: 

colloidal  proteins  ?^  crystalloid  amino  acids 
colloidal  o'lycQcren  5=^  crystalloid  sugar 
nondiffusihlo  fats  ^  diffusible  soaps  and  glycerol. 

In  consequence  of  this,  if  the  crystalloids  dififuse  from  the  blood  into 
a  cell  there  is  at  once  an  excess  of  this  end  of  the  equation,  and, 
hastened  by  the  intracellular  enzjones,  partial  synthesis  to  the  colloid 
soon  occurs  to  establish  chemical  equilibrium.  Chemical  changes  in 
the  crystalloids,  by  oxidation,  reduction  or  hydrolysis,  upset  this  chem- 
ical equilibrium,  and  hence  further  diffusion,  synthesis  and  hydrolysis 
continue,  one  upsetting  the  other  continuously.  If  equilibrium  were 
established  we  should  have  no  further  reactions,  and  the  cells  would 
be  inactive.  The  constant  upsetting  of  the  equilibrium  is  what  con- 
stitutes cell  life. 

The  relation  of  osmotic  pressure  and  osmosis  to  physiological  prob- 
lems is  only  beginning  to  be  studied.  It  is  apparent  that  they  must 
be  of  essential  importance  in  absorption  from  the  alimentary  canal, 
in  absorption  and  excretion  between  the  cells  and  the  blood  stream, 
and  in  secretion  by  glandular  organs;  but  it  is  also  certain  that 
they  are  no  less  important  in  all  the  less  obvious  chemical  and  phys- 
ical processes  of  the  cell.^'^  In  pathological  processes  osmotic  pressure 
may  play  an  equally  important  role,  and  the  facts  discussed  in  the 
preceding  paragraphs  will  be  alluded  to  frequently  in  subsequent 
chapters. 

1'  For   further  consideration  of  the  subject  of  osmotic  pressure  in   these  rela- 
tions see:    Livingston,   "The  Role  of  Diffusion  and  Osmotic  Pressure  in  Plants." 
University  of  Chicaeo  Press,   Cliicago.    100.3;    Czapek.  "Biocheniie  der   Pflanzen," 
Jena,  lfl03.     Also,  Spiro,  Pauli  and  Ilobcr,  all  previously  cited. 
3 


34  TUB    CHEMISTRY    AND    PHYSICS    OF    THE    CELL 

COLLOIDS  18 

Since  Graham  in  1861  studied  the  differences  between  the  sub- 
stances that  did  or  did  not  diffuse  readily  through  animal  or  parch- 
ment membranes,  soluble  substances  have  been  classified  in  the  two 
main  groups  of  colloids  and  crystalloids,  which  distinction  Graham 
believed  separated  two  entirely  different  classes  of  matter.  Although 
at  the  present  time  the  differences  between  the  two  classes  do  not 
seem  so  great,  yet  the  same  division  is  found  useful  in  classification. 
By  colloids  Graham  indicated  those  substances  which  were  dissolved 
to  the  extent  of  showing  no  visible  particles  in  suspension,  but  which 
either  did  not  pass  through  diffusion  membranes  at  all,  or  did  so  very 
si  owl}'  indeed,  as  compared  to  the  crystalloid  substances.  Under  cer- 
tain conditions  they  tended  to  assume  a  sticky,  glue-like  nature, 
hence  the  name.  (Many  substances  are  now  known  which  have  the 
chief  properties  of  the  colloids  and  are  therefore  classified  among 
them,  but  never  are  glue-like,  e.  g.,  the  colloidal  metals,  so  that  the 
name  has  lost  some  of  its  original  significance.)  The  physical  prop- 
erty which  Graham  particularly  noted  in  the  colloids,  besides  their 
non-diffusibility,  was  the  tendency  to  assume  various  states  of  solidity. 
Not  only  can  they  be  in  solution,  when  he  called  them  "sols"  (when 
the  solvent  Avas  water,  "hydrosols"),  but  they  can  become  quite  firm 
although  containing  much  water  (then  called  "gels"  or  "hydrogels"). 
The  gels  may  assume  a  firm,  coagulated  condition,  the  so-called  "pec- 
tous"  state,  which  state  is  permanent  in  that  the  gel  form  cannot  be 
reobtained  from  the  pectous  modification.  Finally  the  colloid  can  be 
in  a  dry,  solid  state,  quite  free  from  water,  and  then  not  a  sol  at  all. 

Included  in  the  great  class  of  colloids  are  all  forms  of  proteins, 
and  also  giims,  starch,  dextrin,  glycogen,  tannin,  chondrin,  probably 
the  enzymes,  and  also  the  greater  number  of  organic  dyes;  also  there 
are  inorganic  colloids,  such  as  silicic  acid,  arsenic  sulphide,  hydrated 
oxide  of  iron,  and  many  other  similar  compounds,  besides  the  ele- 
ments themselves,  especially  the  noble  metals,  which  may  exist  in  col- 
loidal form.  It  will  be  seen  at  once  that  the  chief  constituents  of  the 
cells,  in  fact  nearly  all  the  primaiy  constituents  except  the  inorganic 
salts,  are  organic  colloids,  and  therefore  the  properties  of  the  cells 
are  largely  dependent  upon  the  properties  of  the  colloids. 

In  considering  the  characteristics  of  the  colloids  we  at  once  meet 
the  question — "What  distinguishes  the  colloids  from  the  crystalloids, 
on  the  one  side,  and  from  susjx'nsions  or  emulsions  on  the  other? 
The  sum  and  substance  of  our  present  conception  of  the  nature  of 
colloidal  solution  may  be  briefly  summarized  as  follows: 

1^  For  full  disnissinns  of  tho  nature  of  colloids  soo:  TTolior.  "Physikalisclie 
Clicinio  (Icr  Zcllc."  Loip/,i<r.  1014 ;  Pauli,  Kr<:olinisso  dcr  Pliysiolotrio,  1007  ((>), 
10;");  r.ocliliold,  "Die  Kolloidc  in  Biolo^io  und  :\1(Hlizin,"  Drosdon.  1!)12:  Wo.  Ost- 
wald,  "Crundriss  dor  Kolloidcliomio,"'  Drosdon,  1000;  franslafod  by  'M.  IT.  Fischer, 
101.').  A  good  brief  discussion  of  colloids  is  f!;iven  by  Young  in  Zinsser's  "Infection 
and  Itesistance,"  1014. 


COLLOIDS  35 

It  is  possible  for  solid  substances  to  be  so  divided  among  the  par- 
ticles of  a  solvent  that  they  remain  permanently  in  this  condition, 
neither  aggregating  into  masses  nor  separating  out  through  the  action 
of  gravity.  Witli  some  substances,  as  sugar,  for  example,  the  solid 
seems  to  divide  up  into  its  molecular  form,  eacli  molecule  being  free 
from  all  others  of  its  kind  except  during  occasional  contacts.  Some 
other  substances,  as  salt,  go  still  further,  and  the  molecule  divides  into 
two  or  more  parts,  which  have  different  electric  charges  (ionization) . 
The  first  of  these  classes  of  substances  forms  a  solution  which  con- 
tains no  particles  visible  by  any  known  means,  does  not  contain 
particles  large  enough  to  reflect  light  impinging  upon  them,  exerts 
a  large  osmotic  pressure,  but  does  not  conduct  electricity.  The 
other,  in  which  ionization  has  occurred,  differs  solely  in  its  capacity 
to  conduct  electricity  readily.  Both  are  true  solutions  of  crj-stal- 
loids;  the  one  which  does  not  ionize  is  a  non-electrolijte ;  the  other,  by 
virtue  of  its  ionization,  is  an  electrolyte,  the  ions  carrying  electric 
charges  through  the  solution. 

At  the  other  end  of  the  scale  we  have  substances  which  are  quite 
insoluble  when  in  masses,  but  which,  when  very  finely  divided  by  me- 
chanical means,  can  be  suspended  and  uniformly  distributed  through 
a  fluid  without  having  any  marked  tendency  to  aggregate  or  settle 
out.  Such  suspensions  or  emulsions  contain  particles  visible  under 
the  microscope,  usually  appear  turbid,  refract  light,  are  non-diftusible, 
exert  no  osmotic  pressure,  and  do  not  transmit  electricity.  Such  mix- 
tures are  obviously  very  different  from  the  true  solutions  above  de- 
scribed. Between  these  two  extremes  stand  the  colloids,  which  vary 
in  their  properties  so  that  they  approach  sometimes  the  suspensions 
(e.  g.,  lecitliin,  or'  coagulated  egg-albumin  in  colloidal  suspension), 
and  sometimes  more  nearly  the  true  solutions  (e.  g.,  dextrin).  No 
sharp  boundaries  can  be  drawn  between  any  of  tlie  members  of  the 
series.  Indeed,  one  substance  may  present  all  the  different  stages 
under  different  conditions,  some  agreeing  with  the  properties  of  the 
typical  suspensions,  and  some  with  the  properties  of  the  true  solutions. 
The  colloids  stand  in  an  intermediary  position,  differing  quanti- 
tatively in  one  way  or  another  from  the  true  solutions,  but  yet  ap- 
proaching them  closely  and  sometimes  almost  indistinguishably  re- 
sembling them.  For  the  most  part,  however,  they  show  character- 
istics decided  enough  to  entitle  them  to  separate  classification,  and  to 
make  any  confusion  with  the  crystalloids  impossible. 

The  Characteristics  of  Colloids. — The  chief  properties  of  the 
colloids  are,  then,  as  follows : 

Amorphous  Fcrai. — This,  like  almost  all  other  "colloidal  properties," 
is  not  absolute,  for  in  egg-albumin,  hemoglobin,  and  various  globulins 
w^e  have  proteins  which  in  every  respect  are  typical  colloids,  yet 
they  form  crystals  readily  and  abundantly.  Oxyhemoglobin,  the  mo- 
lecular weight  of  which  is  calculated  at  about  14,000,  exhibits  Tyn- 


36  THE    CHEMISTRY    AHiD    PHYSICS    OF    THE    CELL 

dall's  phenomenon  and  will  not  pass  through  a  very  fine  porcelain 
filter,  and  therefore  resembles  the  colloids  decidedly,  yet  it  forms 
beautiful  crystals.  The  very  fact  that  crystals  are  formed,  Spiro 
points  out,  is  proof  that  when  in  soluti(jn  tlie  individual  molecules 
must  have  been  free  and  separate,  for  otherwise  they  could  scarcely 
unite  in  the  definite  spatial  relations  necessary  to  produce  crystalline 
foi-ms.  With  these  few  exceptions,  however,  the  colloids  do  not  pre- 
sent any  typical  structure,  and  are  not  crystalline  under  any  visible 
condition.  But  when  they  are  made  insoluble  by  chemical  means 
they  may,  under  certain  conditions,  produce  rather  characteristic 
non-crystalline  structures,  a  matter  that  will  be  discussed  in  a  sub- 
sequent ])aragTaph. 

Solubility. — Although  we  speak  of  "colloidal  solutions,"  this  term 
does  not  commit  us  to  the  theory  of  the  identity  of  the  solution  of 
colloids  with  that  of  crystalloids.  We  have  above  stated  w^hat  seems 
to  be  a  fair  view  of  the  matter  as  shown  by  many  methods  of  experi- 
mentation. Most  colloids  seem  to  be,  in  fact,  suspensions  of  masses 
of  molecules,  or  perhaps  of  very  large  sing:le  molecules,  and  a  true 
solution  is  likewise  a  suspension  of  single  molecules  or  of  ions.  When 
the  aggregations  of  molecules  are  sufficiently  large,  w^e  have  an  ordi- 
nary suspension ;  but  a  single  protein  molecule  is  as  large  as  a  very 
great  number  of  molecules  of  such  substances  as  sugar  (crystalloid)  ; 
or  tannin,  C14H10O9  (colloid)  ;  or  calcium  carbonate  (insoluble,  sus- 
pension) ;  and  it  would  be  strange  if  a  true  solution  of  a  protein 
did  not  behave  in  many  particulars  like  a  suspension  of  molecular 
aggregates  of  dimensions  similar  to  the  dimensions  of  protein  mole- 
cules. Nearly  all  colloidal  solutions  show  Tyndall's  phenomenon, 
which  demonstrates  the  existence  of  particles  in  suspension  large 
enough  to  reflect  light  from  their  surfaces.  Most  of  the  colloids  are 
held  back  by  very  fine  filters  to  a  greater  or  less  degree ;  some  are  al- 
most entirely  retained  by  a  hardened  paper  filter,  while  others  pass 
through  the  finest-pored  clay  filters.  Furthermore,  the  metallic  col- 
loids, such  as  those  of  platinum,  gold,  and  silver,  are  unquestionably 
suspensions  of  finely  divided  particles  of  metal,  yet  they  exhibit  all  the 
typical  phenomena  of  colloids,  passing  through  many  sorts  of  filtei-s, 
and  even  accomplishing  the  same  hydrolytic  changes  as  many  en- 
zymes. 

It  must  also  be  mentioned  that  the  solvent  is  probably  an  im- 
portant factor  in  determining  tlie  colloidal  or  noii-colloidal  nature 
of  a  substance ;  e.  g.,  soaps  form  true  solutions  in  alcoliol  and  colloidal 
solutions  in  water;  gelatin  forms  colloidal  solutions  in  water  but  not 
in  ether,  whereas  rubber  forms  colloidal  solutions  in  ether  but  not  in 
water. 

Closely  I'elati'd  to  solubility  is  tiie  ])li('ii(itiieii()n  ol'  iuihihittnn  (the 
"Quellung"  of  German  writers),  which   inny   he  deliuiMl   ;is  the  tak- 


COLLOIDS  37 

inp:  up  of  a  fluid  hy  a  solid  body  witliout  chemical  change.  Not 
all  colloids  possess  this  i)roperty,  but  it  is  shown  by  most  of  the 
organic  colloids,  particularly  the  proteins.  Fick  distinguishes  cap- 
illary, osmotic,  and  molecular  imbibition,  the  latter  of  which  is  the 
form  exhibited  by  colloids,  and  it  occurs  independent  of  the  existence 
of  pores  or  other  preformed  spaces  in  the  imbibing  body.  The  imbibi- 
tion of  water  by  colloids  is  more  than  a  simple  mechanical  process, 
for  it  is  accompanied  by  a  contraction  in  the  total  volume  of  solid 
iiiid  water,  and  by  the  evolution  of  lieat.  The  forces  developed  are 
far  greater  than  those  of  osmotic  pressure ;  e.  g.,  to  prevent  imbibi- 
tion of  water  by  starch  requires  a  pressure  of  over  2500  atmospheres. 
On  the  other  hand,  the  ]^hysical  properties  of  an  aqueous  colloidal  so- 
lution show  that  the  colloid  is  not  chemically  combined  in  the  form 
of  a  hydrate.  To  describe  this  peculiar  relation  Hofmeister  and  Os- 
wald recommend  the  term  "mechanical  affinity."  Hardy  has  shown 
that  water  held  in  a  gelatin  jelly  cannot  be  removed  by  g-reat  pres- 
sures (400  pounds  to  the  square  inch),  but  after  the  nature  of  the 
jelly  is  so  changed  by  formalin  that  it  is  no  longer  liquefiable  by  heat, 
the  water  can  be  easily  expressed  from  the  loose  meshwork  that  is 
formed.  It  would  seem  from  this  that  the  imbibition  and  retention 
of  water  by  colloids  may  be  closely  related  to  surface  phenomena. 
Hofmeister  has  shown  that  organized  animal  tissues  obey  the  same 
laws  of  imbibition  as  do  simple  gelatin  plates,  and  probably  this  phe- 
nomenon of  colloids  is  very  important  in  physiological  and  patho- 
logical processes. 

Non-diffusibility. — The  lack  of  power  to  pass  through  animal  and 
parchment  membranes,  wiiich  was  Graham's  starting-point  in  the 
study  of  colloids,  is  also  only  a  relative  condition.  This  is  shown  by 
the  following  figures,  giving  the  relative  time  required  by  the  same 
amount  of  different  substances  to  pass  through  a  certain  diffusion 
membrane : 

Sodium  chloride 2.3.3 

Sugar 7.00 

Magnesium  sulphate 7.00 

Protein 49.00 

Caramel 98.00 

This  difference  of  time  is  so  great,  however,  as  to  pennit  of  separation 
of  salts  from  proteins,  etc.,  by  dialyzation,  a  process  in  constant  u.se. 
Primarily  the  ability  to  diffuse  through  a  given  membrane  requires 
that  the  diffusing  substance  be  soluble  in  the  membrane.  Diffusion 
membranes  are  always  composed  of  colloids,  e.  g.,  animal  bladders,  or 
parchment,  which  is  a  colloidal  cellulose.  Crystalloids  are  generally 
soluble  in  colloids,  while  colloids  are  little  or  not  at  all  soluble  in 
other  colloids,  and  hence  do  not  diffuse  through  one  another  readily 
and  permeate   diffusion   membranes  very  slowly.     For  example,    if 


38  THE    CHEMISTRY    AND    PHYSICS    OF    THE    CELL 

a  stick  of  agar  jelly  be  placed  in  a  solution  of  aminoniated  cop- 
per sulphate  (a  crystalloid),  and  another  be  placed  in  a  solution  of 
Prussian  blue  (a  colloid),  it  will  be  found  that  the  copper  solution 
penetrates  the  agar  completely  before  the  colloidal  solution  of  Prus- 
sian blue  has  penetrated  it  at  all.  This  property  is  of  great  im- 
portance, undoubtedly,  in  keeping  different  colloidal  constituents  of 
the  cell  in  given  localities  within  its  protoplasm,  e.  g.,  the  oxidizing 
ferments  seem  to  be  chiefly  localized  within  the  nucleus;  the  colloidal 
glj'cogen  remains  where  it  is  formed  in  the  cytoplasm,  unable  to 
escape  from  the  cell,  whereas  the  crystalloidal  sugar  from  which  it  is 
formed  and  into  which  it  is  converted,  dififuses  rapidlj'  into  or  out  of 
the  cell. 

The  osmotic  pressure  of  the  colloids  is  extremely  small.  The  closely 
related  phenomena  of  diffusion,  depression  of  freezing -point,  and  ele- 
vation of  hoiUng-point,  are  also  exhibited  by  colloids  to  but  an  ex- 
tremely slight  degree.  For  example,  in  one  experiment,  the  dissolv- 
ing of  from  14  per  cent,  to  44  per  cent,  of  egg-albumin  in  water  low- 
ered the  freezing-point  but  0.02°  to  0.06° ;  and  some  other  colloids 
have  even  less  effect.  The  results  of  the  latest  and  best  experiments 
seem  to  indicate  that  the  trifling  effects  of  colloids  upon  osmotic  pres- 
sure and  upon  freezing-  and  boiling-points  observed  in  colloidal  solu- 
tions are  due  to  the  colloids  themselves,  rather  than  to  included  im- 
purities, although  it  may  possibly  be  that  some  of  these  effects  are 
due  to  the  high  surface  tension  and  cohesion  affinity  of  the  colloids. 
In  all  cellular  processes  accompanied  by  manifestations  of  osmotic 
pressure  or  dififusion,  however,  the  crystalloids  may  be  considered 
as  almost  entirely  responsible. 

Electrical  Phenomena. — As  colloids  do  not  separate  freely  into  ions 
when  dissolved,  they  do  not  conduct  electricity  appreciably.  How- 
ever, when  an  electric  current  is  passed  through  water  containing 
■colloids  in  solution,  the  colloidal  particles  tend  to  pass  to  one  pole  or 
the  other.  Most  colloids  move  toward  the  anode.  This  phenomenon. 
eataphoresis,  is  also  generally  exhibited  by  suspensions,  and  hence  in 
this  particular  the  colloids  resemble  suspensions  rather  than  solutions. 
Helmholtz  has  explained  the  movement  of  the  suspended  particles  as 
due  to  the  accunmlation  of  electrical  charges  u])on  the  surfaces  of  two 
heterogeneous  media  when  in  contact.  The  nature  of  the  charge  de- 
pends upon  both  the  suspended  substance  and  the  fluid ;  e.  g.,  sulphur 
or  graphite  particles  suspended  in  water  assume  a  negative  charge 
and  move  toward  the  anode,  but  when  suspended  in  oil  of  turpentine 
they  become  positively  charged  and  move  toward  the  cathode.  AYater 
has  such  a  high  dielectric  constant  that  most  substances  suspended  in 
water  become  negatively  charged  as  eoni])ared  with  the  water,  and 
move  toward  the  positive  pole  or  anode. 

Hardy  has  observed  that  colloidal  solutions  of  coagulated  proteins 
move  toward   the   anode  when    in   alkaline  solnlion,   and   toward    the 


COLLOIDS  39 

cathode  when  in  acid  solution.^"  Tliis  peculiar  property  of  proteins 
sugorests  that  perhaps  simple  surface  phenomena  do  not  snfifice  to  ac- 
count for  the  electrification  of  all  colloid  particles.  Knowing  the  pe- 
culiar amphoteric  character  of  ])r()tcins,  which  is  probably  due  to  the 
presence  of  both  NII2  and  COOII  groups  in  the  molecule,  we  can 
readily  understand  that  in  an  acid  solution  the  NH2  radicles  are  com- 
bined with  the  acid,  leaving  the  COOH  iradicles  free.  The  molecule 
would  then  have  acid  properties,  and  could  dissociate  into  an  acid  II 
ion  and  a  basic  or  electrically  positive  colloid  ion.  The  colloid  ion 
would  then  go  toward  the  negative  pole  slowly,  because  of  its  great 
size.  When  a  suitable  concentration  of  both  ions  is  produced  the  pro- 
teins will  move  towards  both  poles,  this  concentration  being,  in  the 
case  of  serum  albumin.  H^IO  "  (^lichaelis).  Living  protoplasm  be- 
haves in  most  instances,  as  if  the  proteins  were  acids  bound  to  inor- 
ganic cations  (Robertson),  and  is  usually  stimulated  at  the  cathode 
on  the  '"make"  of  the  current.  It  is  permeable  to  ions,  and  the 
vitality  of  a  tissue  is  so  dependent  on  the  maintenance  of  normal 
permeability  that  the  permeability  may  be  employed  as  a  sensitive 
and  reliable  indicator  of  its  vitality  (Osterhout  ^°^).  This  may  be 
done  by  determining  the  electrical  resistance  of  the  cells,  which  is 
lowered  by  anything  that  lowers  their  vitality. 

Surface  tension,-"  which  may  be  described  as  the  force  icith  tchich  a 
fluid  is  strii'i)ig  to  reduce  its  free  surface  to  a  minimum,  is  highly 
exhibited  by  colloids  as  compared  with  crystalloids.  The  formation  of 
emulsions  and  the  spreading  out  of  oil  upon  the  surface  of  water  de- 
pend upon  surface  tension.  Ameboid  movement  may  be  attributed  to 
changes  in  surface  tension,  as  also  may  phagocytosis.  (The  relation 
of  surface  tension  to  these  processes  will  be  considered  under  the 
subject  of  Inflammation.) 

The  effect  of  colloids  upon  chemical  processes  going  on  within 
their  solutions  or  gels  is  surprisingly  small.  Salts  in  solution  in  a 
thick  gel  of  agar  or  gelatin  will  diffuse  almost  as  rapidly  as  in  water ; 
they  will  also  ionize  as  rapidly  as  in  watery  solutions,  and  chemical 
reactions  occur  with  nearly  the  same  speed  and  completeness  as  if  the 
colloids  were  absent.  Furthermore  it  makes  little  difference  whether 
these  processes  are  measured  in  a  colloid  solution  that  is  liquid,  or 
after  it  has  set  in  the  gel  form.  These  facts  merely  indicate  that  the 
colloids  do  not  greatly  impede  the  movements  of  molecules  or  ions  in 
solutions.  On  the  other  hand,  as  before  mentioned,  colloids  diffuse 
very  slowly  into  each  other.  Hence,  in  the  cell  the  colloids  are  quite 
fixed  in  their  positions,  whereas  the  crystalloids  may  wander  about 
freely,  and  this  arrangement  is  certainly  of  great  importance  in  bio- 

19  According  to  Field  and  Teague  (Jour.  Exper.  Med.,  1007  (9),  222).  vative 
proteins  in  serum  move  towards  the  cathode,  no  matter  what  the  reaction. 

ina  Science.  1904    (40),  4SS. 

20  See  article  on  "Surface  Tension  and  Vital  Phenomena."  hv  ^Macallum.  Erirel)- 
nisse  d.  Phvsiol.,  1011    (11).  (\()2. 


40  Tin:  CHEMISTRY  Ayo  riivfiics  of  the  cell 

logic  processes.  Pauli  suggests  the  probability  that  the  fixation  of  the 
colloid  causes  the  cell  to  have  different  properties  in  different  parts, 
and  so  various  reactions  may  occur  independently  in  different  areas 
of  the  cytoplasm.  The  possibility  of  the  correctness  of  this  view  is 
increased  when  we  consider  that  the  enzymes  are  colloids,  for  there  is 
inuch  evidence  to  show  that  they  are  distributed  in  just  such  an  un- 
even manner  within  the  cells. 

Although  colloids  permit  the  passage  of  dissolved  crystalloids, 
through  them,  they  greatly  interfere  with  the  movement  of  larger 
particles.  This  property  accounts  for  the  ability  of  colloids  to  hold 
many  insoluble  substances  in  such  extremely  fine  suspensions  that 
they  seem  superficially  to  be  in  true  solution.  If,  for  example,  sodium 
phosphate  is  added  to  a  solution  of  casein  in  lime-water,  tlie  calcium 
phosphate  formed  does  not  precipitate.  It  is  not  in  solution,  how- 
ever, but  rather  exists  as  a  suspension  of  very  finely  divided  particles 
of  the  salt  which  the  colloid  keeps  from  aggregating  into  particles, 
large  enough  to  be  visible  or  to  overcome  the  viscosity  of  the  fluid 
and  sink  to  the  bottom.  Probably  in  this  way  many  substances,  in- 
cluding calcium  salts,  are  carried  in  the  blood,  held  in  permanent 
suspension  b3'  the  proteins.  Substances  thus  finely  divided  will  have 
extremely  large  surface  area  for  reactions,  and,  therefore,  will  un- 
doubtedly undergo  changes  with  considerable  rapidity  and  facility, 
although  not  in  solution. 

Precipitation  and  Coagulation  of  Colloids. — Because  of  the  rather 
slender  marghi  by  which  the  colloids  are  separated  from  the  suspen- 
sions, their  persistence  in  solution  is  generally  in  a  rather  precarious 
condition.  Kelatively  slight  changes  suffice  to  throw  the  colloids  out 
of  solution,  and  when  once  precipitated,  they  are  often  incapable  of 
again  dissolving  in  the  same  solvent.  Solutions  of  albumin  may  un- 
dergo spontaneous  coagulation  on  standing  for  some  time,  and  agita- 
tion rapidly  produces  the  same  effect  in  many  protein  solutions. 
Some  inorganic  colloids  are  as  readily  coagulated  as  the  proteins. 
A  comparatively  small  rise  in  temperature,  less  than  to  50°  C.  with 
some  proteins,  renders  the  protein  perfectly  insoluble.  Further- 
more, we  have  coagulation  of  protein  solutions  by  enzyme  action. 
The  inorganic  "colloidal  suspensions"  may  be  precipitated  by  the 
addition  of  very  small  quantities  of  electrolytes.  Colloidal  solutions 
of  the  type  of  the  proteins  are  not  so  readily  i)recipitated  by  most 
electrolytes,  but  if  to  the  solution  large  quantities  of  crystalloids  are 
added,  the  protein  molecules  are  practically  crowded  out  of  solution, 
as  in  the  "salting-out"  process  used  in  separating  proteins  by  am- 
monium sul])liate  and  other  salts.  Tlie  effect  of  heat  U])on  different 
folloids  is  ])ecu]iar,  in  that  some  vai'ieties,  a.s  silicic  acid,  aluminium 
liydrate,  and  many  proteins  are  rendcrcM]  so  insolnble  tliat  they  can- 
not again  be  dissolved  in  any  fluid  williont  lii-st  l)eing  modified  in 
some  way;  whereas  colloids  of  the  type  of  gelatin  and  agar  are  made- 


COLLOIDS  41 

more  soluble  by  heat.  The  change  of  colloids  into  insoluble  forms, 
the  'Specious''  condition  of  (irahani,  re(|uir('.s  the  presence  of  water, 
for  the  dry  colloids  ni;i\  lie  iieated  to  relatively  hiprh  temperatures 
witliout  losino-  their  s()lul)ilit y.  On  the  other  hand,  deliydration  of 
colloids  while  in  solution  will  result  in  their  precipitation  and  coagu- 
lation, as  occurs,  in  protein  solutions  when  alcohol  is  added. 

If  solutions  of  two  oppositely  char<ied  colloids  are  broujrht  together 
they  nuiy  ])recipitate,  but  if  either  is  present  in  excess  the  precii)ita- 
tion  may  be  incomplete,  or  even  completely  absent.  This  inhibition 
of  precipitation  is  of  particular  interest  because  it  so  closely  resem- 
bles the  phenomenon  observed  in  the  precipitin  reaction,  whereby  an 
excess  of  the  antigenic  protein  will  ])revent  precipitation.  Also  cer- 
tain colloids  will  prevent  the  precipitation  of  other  colloids  by  elec- 
trolytes, which  fact  is  tlie  basis  of  the  Lange  reaction  of  spinal  fluid 
with  colloidal  gold. 

Colloids  are  precipitated  by  many  electrolytes,  apparently  through 
the  formation  of  true  ion  compounds,  one  or  both  of  the  ions  of  the 
electrolytes  uniting  with  the  colloid  ion :  although  some  writers,  as 
Spiro,  believe  that  the  combination  is  merely  an  additive  one  between 
entire  molecules.  ^Mathews  -^  has  advanced  the  theory  that  the  solu- 
tion tension  of  the  ions  is  an  important  factor  in  determining  the  pre- 
cipitation of  colloids  by  electrolytes.  In  general,  precipitation  of 
colloids  results  from  the  reduction  of  the  surface  in  proportion  to  the 
mass,  because  of  an  aggregation  of  the  particles ;  this  may  be  brought 
about  by  changing  the  surface  electrical  conditions,  by  uniting  the 
molecules  chemically,  or  by  reducing  the  amount  of  the  solvent. 

The  Structure  of  Colloids  and  of  Protoplasm. — Two  very  differ- 
ent sorts  of  substances  are  usually  included  under  the  term  colloid, 
because  they  show  the  essential  features  of  colloids  in  most  respects; 
but  as  in  many  other  respects  they  are  quite  unlike  each  other,  it  may 
be  well  to  distinguish  between  them  in  some  way.  As  a  type  of  one 
class  we  may  take  gelatin ;  of  the  other,  such  a  substance  as  colloidal 
arsenious  sulphide.  Gelatin  solutions  form  gels  upon  cooling  or  evap- 
oration, and  redissolve  when  heated  or  when  more  solvent  is  added. 
Arsenious  sulphide  does  not  form  gels  upon  cooling,  and  when  solidified 
in  any  way,  does  not  redissolve.  In  addition,  the  gelatin  type  is  very 
viscous,  and  is  not  coagulated  by  the  presence  of  salts  unless  these  are 
added  in  large  amounts;  while  the  other  type  does  not  render  the 
fluid  in  which  it  is  dissolved  appreciably  more  viscid,  and  it  forms  a 
precipitate  immediately  if  minute  amounts  of  electrolytes  are  intro- 
duced. As  the  former  type  resembles  in  many  details  the  true  solu- 
tions, while  the  latter  approaches  more  closely  to  the  suspensions,  it 
has  been  proposed  to  distinguish  them  by  the  terms  "colloidal  so- 
lution" and  "colloidal  suspension."  --     Of  the  two  types,  the  colloidal 

21  American  Journal  of  Phvsiolofry.  190.5    (14),  203. 
22^0768,  Amorican  Chemical  Journal,  100.5    (27),  85. 


42  THE    CHEMISTRY    AXD    PHYSICS    OF    THE    CELL 

solutions  are  by  far  the  more  important  in  l)iological  considerations, 
since  the  colloidal  suspensions  are  usuall}"  prepared  artifieiaUy  and 
seldom  occur  in  nature,  e.  g.,  Bredig's  colloidal  suspensions  of  the 
noble  metals. 

The  colloidal  solutions  of  proteins,  which  constitute  the  chief 
part  of  every  cell,  are  of  two  types — one,  such  as  albumin,  forms 
a  coagulum  when  heated,  which  under  ordinary^  conditions  is  not 
reversible ;  that  is,  it  does  not  again  go  into  solution.  Gelatin,  how- 
ever, becomes  more  fluid  when  heated,  and  when  cooled,  it  forms 
a  gel  which  is  readily  reversible  to  the  soluble  form  under  the  influ- 
ence of  heat.  Agar  is  another  familiar  example  of  this  heat-reversible 
type.  Within  the  cell,  so  far  as  we  know,  occur  only  the  first  type, 
the  proteins  that  form  non-reversible  coagula. 

An  extensive  study  of  the  physical  structure  of  the  colloids  has 
been  made  by  Hardy.-^  As  long  as  the  colloid  is  in  solution  it  is 
structureless,  although,  as  before  mentioned,  the  existence  of  free 
solid  particles  can  be  demonstrated  by  certain  optical  methods.  The 
solution  is  homogeneous,  and  although  perhaps  viscid,  still  it  is  a  typ- 
ical solution.  Such  solutions  can  become  solid,  either  by  the  effect  of 
temperature,  of  certain  chemical  fixing  agents,  or  physical  means. 
It  was  found  h\  Hardy  that  in  undergoing  this  solidification  there  oc- 
curred a  separation  of  the  solid  from  the  liquid,  the  solid  particles 
adhering  to  form  a  framework  holding  the  liquid  within  its  intei^tices. 
Heat-reversible  gels  show  no  structure  until  they  are  made  irreversi- 
ble by  hardening  agents,  etc. ;  e.  g.,  a  jelly  of  gelatin  appears  struc- 
tureless, but  when  treated  with  formalin  or  other  fixing  agent,  the 
structural  appearances  described  below  appear.  The  figures  formed 
by  the  framework  vary  according  to  the  nature  and  concentration 
of  the  colloid  and  of  the  solvent,  and  also  with  the  fixing  agent  used, 
the  temperature,  and  the  presence  or  absence  of  extraneous  substances. 
In  general,  however,  the  figures  obtained  in  the  solidification  of  protein 
solutions  by  fixing  agents,  such  as  bichloride  of  mercury  or  formalin, 
hear  a  striking  reseniblance  to  the  finer  structures  of  protoplasm  as 
described  by  cytologists.  There  is  produced  an  open  network  struc- 
ture with  spherical  masses  at  the  nodal  points,  or  minute  vesicles  hol- 
lowed out  in  a  solid  mass,  or  a  honeycomb  appearance,  or,  when  the 
concentration  of  the  colloid  is  very  slight,  perhaps  there  is  only  a 
precipitation  of  fine  granules  of  protein  such  as  we  often  see  in  histo- 
logical preparations  of  edematous  cells  and  tissues.  All  these  forms 
seem  to  depend  cliiefly  u])on  the  concentration  of  the  colloid.  Tlie 
important  fact  is  that  when  the  chemicals  ordinarily  used  as  fixatives 
of  cells  for  histological  purposes  act  upon  solutions  of  colloids  that 
are  perfectly  homogeneous,  they  produce  very  constant  and  charac- 
teristic formations  wliicli  recall  at  once  the  structures  found  in  the 
protoplasm  of  hai-deiied  cells.     IMoreover,  the  use  of  different  fixing 

23.|,,nriial   nf  riiysiolo^ry,   ISOO    (2-1).   loS. 


STRUCTURE  OF  CELLS  43 

agents,  such  as  osinic  acid,  formalin,  and  liidiloride  of  mercury,  pro- 
duces just  the  same  differences  in  the  structure  of  colloidal  solutions 
that  they  i)roduce  in  the  protoplasm  of  cells  hardened  by  them. 
Neither  are  the  appearances  seen  in  unfixed  specimens  reliable  indi- 
cations of  the  true  structure  of  the  living  protoplasm.  Granules  of 
secretion  may  disappear  after  or  during  the  death  of  the  cell  (e.  g., 
glycogen)  or  they  may  swell  up  [e.  <j.,  nnu'in  graiuiles),  thus  giving 
the  appearance  of  a  network  or  honeycomb  which  is  then  incorrectly 
ascribed  to  the  protoplasm  itself.  Death  of  the  cells,  even  when  not 
produced  by  external  influences,  seems  to  be  accompanied  by  coagula- 
tion of  some  parts  of  the  cell  constituents,  and  hence  a  cell  examined 
in  anj'thing  but  its  normal  living  condition,  an  extremely  difficult 
matter,  will  not  present  a  true  idea  of  how  it  appears  and  is  composed 
while  in  that  condition. 

If,  with  these  facts  in  mind,  we  consider  the  theories  of  morpholo- 
gists  as  to  the  finer  structure  of  the  cell  protoplasm  based  upon  stud- 
ies of  cells  fixed  in  various  hardening  agents,  it  becomes  evident  that 
the  possibility  that  the  "foam  structure"  advocated  by  Riitschli,  or 
the  "thread,"  "reticular,"  and  "pseudo-alveolar"  structures  of  Fro- 
mann,  Arnold,  Reinke,  and  others,  are  all  simply  the  effect  of  fixatives 
upon  colloid  solutions,  is  very  real.  The  objection  always  advanced 
to  these  theories  of  protoplasmic  structure,  namely,  that  the  struc- 
tures described  were  artificial  productions,  not  present  in  the  normal 
living  cell,  and  variously  described  and  interpreted  by  differ- 
ent investigators,  because  each  worked  with  a  different  hardening 
fluid  or  different  technic,  is  strongly  supported  by  these  observations 
upon  colloids.  The  possibility  that  the  living  protoplasm  is  homo- 
geneous still  remains  open.  This  matter  will  receive  further  consid- 
eration in  the  next  section. 


THE  STRUCTURE  OF  THE  CELL  IN  RELATION  TO  ITS  CHEMIS- 
TRY AND  PHYSICS  -^ 

It  is  obviously  impossible  to  separate  nuclei,  nucleoli,  cytoplasm, 
and  cell  membranes  from  each  other  (except  with  sperm  heads)  and 
to  isolate  them  in  quantities  sufficient  for  analysis,  and  therefore  we 
are  still  quite  uncertain  as  to  just  the  chemical  differences  that  exist 
between  them.  That  there  are  differences  is  certain,  and  by  means  of 
micro-chemical  reactions,  by  comparing  analyses  of  cells  in  which  nu- 
e'eus  or  cytoplasm  predominate,  and  by  stud^ying  their  physico-chem- 
ical relations  to  one  another,  we  have  arrived  at  more  or  less  tangible 
ideas  on  the  question  of  the  relation  of  the  structural  elements  of  the 
cell  to  its  composition. 

23a  Reviews  of  tlio  siffnificanoo  of  cell  struetiirp  for  patholofry  arc  pixcii  by 
Benda  and  Ernst  in  Zentrlbl.  allp:.  Path.,  1014.  P>d.  20.  Erfriinzunpslipft. 


44  nil-:  vhemistry  a\j>  I'livsics  of  the  cell 

THE  NUCLEUS  2* 

Although  the  luieleus  i)reseiits  morphologically  a  sharp  isolation 
from  the  cytoplasm,  and  displays  eciually  sharp  tinctorial  ditt'erences, 
it  is  probable  that  chemically  the  ditt'erences  between  nucleus  and  cy- 
toplasm are  quantitative  rather  than  qualitative.  The  characteristic 
affinit}'  of  certain  elements  of  the  nucleus  for  basic  stains  depends 
upon  the  presence  in  the  nucleus  of  nucleoproteins  in  large  proportion, 
and  to  a  limited  degree  nucleoproteins  are  characteristic  of  nuclei. 
Their  affinity  for  basic  dyes  depends  upon  the  nucleic  acid  radical.-*'* 
For  exami)le,  the  heads  of  spermatozoa  contain  nucleic  acid  bound  to 
simple  proteins  in  such  a  way  that  it  readily  forms  a  salt  or  salt-like 
combination  with  basic  dyes,  and  so  the  sperm  heads  appear  intensely 
stained  by  alum-hematoxylin,  etc.  Ordinary  chromatin  threads  of 
nuclei  appear  to  contain  somewhat  more  firmly  bound  protein  in  their 
nucleoprotein  molecules,  and  hence  stain  less  intensely  than  do  the 
spermatozoa  heads,  except  when  in  karyokinesis,  when  the  chromatin 
nucleoprotein  seems  to  approach  that  of  the  spermatozoa  in  avidity  for 
basic  dyes.  We  also  have  nucleoproteins  with  the  nucleic  acid  so  thor- 
oughly saturated  by  protein  that  they  do  not  stain  at  all  by  basic  dyes, 
and  these  seem  to  exist  principally  in  the  cytoplasm,  and  also  to  form 
the  ground-substance  of  the  nuclei,  occupying  the  spaces  between  the 
chromatin  particles  (this  achromatic  substance  of  the  nuclei  is  called 
linin  or  plastin  by  some  cytologists).  Besides  the  chromatin  and  the 
nucleoli,  there  is  a  peculiar  chromatophile  substance,  suspended  in  the 
liner  part  of  the  nuclear  structure  in  the  same  manner  as  the  chromatin 
itself  is  in  the  coarser  portions ;  this  was  called  lanthanin  by  Heiden- 
hain,-^  and  is  probably  similar  to  the  substances  also  described  as  para- 
ehromatin  and  paraUnin.  Undoubtedly  the  other  forais  of  proteins 
found  in  the  cell,  such  as  globulin,  albumin,  and  nucleoalbumin,  exist 
both  in  the  nucleoplasm  and  in  the  cytoplasm,  the  essential  difference 
being  that  the  proportion  of  nucleoprotein  is  much  greater  in  the  nu- 
cleus. As  nucleoproteins  are  little  affected  by  peptic  digestion,  it  is 
possible  to  isolate  nuclear  elements,  especially  the  chromatin,  for  analy- 
tic purposes,  and  it  has  been  demonstrated  by  this  means  also  that 
nuclein  is  the  chief  constituent  of  the  staining  elements.  The  distribu- 
tion in  the  nucleus,  of  the  other  primary  constituents  of  the  cyto- 
plasm, such  as  lecithin,  cholesterol,  and  inorganic  salts  has  not  yet  been 
worked  out,  except  that  IMacallum  -*'  has  found  that  nuclei  contain  no 
chloi-ide,  as  indicated  by  their  not  staining  with  silver  nitrate,  and 

2^  Karlicr  litcraturo  by  Alhroclit,  "Pathologio  der  Zcllc."  Luliarscli-Ostortafr. 
Erjrcl).  <lcr  allff.  Pathol.,  I'snO  (0),  1000:  see  also  Kossel,  Miiufli.  med.  Wocli..  1011 
(58),  05. 

:;-iH  Tferwerden  (Arch.  Zcllforscli..  l!ll:!  (]'.)).  -l.'il  )  found  lliat  llie  liasopliilic 
firanules  are  disintc^rralcd  spccidcallv  l)y  mudcase.  siijiiiortiiij:  1hi>  view  that  they 
are  mi(deic  acid  com  pounds. 

■■^■-  Fcstsdir.  f.  K.illikcr.   1802,  p.  128. 

-'•i  I'rocecdinL's  of  Ihc  K'oval  Societv.  IHO.-)    (Tfi).  217. 


COMl'OSHT/OX  OF  MCLEI  45 

•<ils(»   no   polassiuin,-'   hut   tlic  ehromatiii   coiitaiiis   lii-iiily   lioiind    ii-oii. 

Nucleoli,  wiiicli  not  all  x'ai'iotios  of  iiudri  |)oss('ss,  diU'ci-  i'fom  Ihe 
otluu'  imeli'ar  .sti'ucturrs  in  liavin*^'  an  at'finily  for  acid  rallicr  tlian  for 
basic  dyes,-"*  at  least  in  fixed  tissues.  Their  clieinical  composition  has 
not  been  ascertained.  Zacliarias  considers  the  nucleoli  as  composed 
of  nuclein  well  saturated  with  protein,  because  of  its  staining  re- 
actions and  its  relative  insolubility  in  alkalies,  and  classes  it  with 
j)lastin  or  linin,  which  forms  the  achromatic  part  of  the  nucleus  and  is 
also  present  in  the  cytoplasm.  jMacallum  -°  found  that  they  reacted 
for  organic  phosphorus  microchemically,  but  less  strongly  than  did 
chromatin  fibers. 

The  nuclear  membrane  is  an  uncertain  structure,  at  times  dense 
and  staining  as  if  formed  of  a  layer  of  chromatin,  in  other  cells  stain- 
ing like  the  cytoplasm  with  which  it  seems  to  be  continuous,  in  most 
cells  disappearing  during  karyokinesis,  and  in  some  protozoa  being 
entirely  absent.  Naturally  the  composition  of  the  nuclear  membrane 
is  unknown,  l)ut  it  is  probable  that  it  acts  as  a  diffusion  membrane  of 
partially  seaiii^ermeable  character,  maintaining  different  conditions 
in  nucleus  and  cytoplasm. 

Functionally  the  nucleus  is  the  essential  element  of  the  cell ;  an  iso- 
lated nucleus  with  but  a  minimum  of  cytoplasm  may  be  able  to  de- 
velop new  c3'toplasm.  Init  isolated  cytoplasm  soon  disintegrates,  al- 
though it  may  manifest  life  for  some  time  by  movement  and  chemical 
activities.  A  popular  theory  is  that  synthetic,  constructive  processes 
occur  in  the  nucleus  or  under  the  influence  of  its  products,  but  to 
what  the  nucleus  owes  these  hypothetical  powers  is  unexplained. 
More  tangible  are  the  theories  based  upon  the  work  of  Spitzer,  Loeb,^° 
Lillie  "^  and  others  which  suggest  that  the  oxidative  processes  of  the 
cell  depend  upon  the  nucleus,  hence  portions  of  the  cell  cut  away  from 
the  nucleus  undergo  asphyxiation.  As  Loeb-says,  "By  cellular  struc- 
ture we  understand  the  fact  that  there  must  be  a  definite  maxinml  dis- 
tance between  the  elements  of  the  protoplasm  and  the  nearest  nu- 
cleus. ' '  However,  more  recent  work  casts  doubt  on  the  dependence  of 
oxidation  on  the  nucleus.'^" 

It  should  be  mentioned  that  certain  cells,  such  as  bacteria  and  algfe, 
seem  to  have  no  true  nuclei,  but  IMacallum ''-  found  that  the  forms  he 
examined  gave  reactions  for  phosphorus  and  iron  in  a  similar  way 
to  the  nucleojiroteins  of  a  nucleus,  suggesting  that  in  such  cells  the 
nuclear  elements  are  diffused  through  the  cell  rather  than  differen- 
tiated.    To    quote    Wilson:     "The    term    'nucleus'    and    'cell    body' 

27  .Jour,  of  Physiol.,  1!)05   (32),  95. 

28  Xvicleoli  of  nerve-cells  are  an  exception,  beinfr  basophilic. 
20Proc.  of  the  Royal  Society,  1S98    (6.3),  M'u . 

30  "Studies  in  General  Physiolntjy,"  Chicago.  190.5. 

31  American  .Journal  of  Plivsiolopv,   1902    (7),  412. 
siaSee  Lillie,  Jour.  Biol.  Ciieni.,  1913   (15),  237. 

32  "Studies  from  the  University  of  Toronto,"  1900. 


46  THE    CHEMISTRY    AND    PHYSICS    OF    THE    CELL 

should  probably  be  regarded  as  only  topographical  expressions,  de- 
noting two  differentiated  areas  in  a  common  structural  basis." 

Because  of  the  relative  acidity  of  the  nuclei  they  are  electrically 
negative  to  the  cytoplasm,  particularly  when  in  karyokinesis,  and  the 
chromatic  elements  of  the  nucleus  can  be  showTi  to  carry  a  negative 
electric  charge.^^  Sperm-heads  in  isotonic  cane-sugar  solution  move 
rapidly — 2000  microns  a  minute — toward  the  anode,  when  a  current 
is  passed  tlirough  the  solution ;  and  leucocytes  also  go  toward  the 
anode  under  the  same  conditions,  the  rate  depending  upon  the  pro- 
portion of  nucleoplasm  and  cytoplasm,  large  leucocytes  sometimes 
even  going  slowly  toward  the  cathode.  The  Sertoli  cells  of  the  testi- 
cle, which  have  a  round  mass  of  cytoplasm  with  a  number  of  minia- 
ture spermatozoa  heads  at  one  side,  orient  themselves  in  the  current 
so  that  the  side  or  end  containing  the  spermatozoa  drags  the  mass 
of  cytoplasm  toward  the  positive  pole. 

THE  CYTOPLASM 

The  cytoplasm,  as  before  mentioned,  contains  all  of  the  primary  cel- 
lular constituents,  and  also  such  secondarj^  constituents  as  the  particu- 
lar cell  possesses.  Nucleoproteins  are  undoubtedly  present  in  unknown 
proportions,  but  with  the  nucleic  acid  well  saturated  by  proteins,  and 
perhaps  also  to  a  large  extent  combined  with  carbohydrates  to  form 
the  glyconucleoproteins.  Sometimes  the  nucleoproteins  o'f  the  cyto- 
plasm may  be  partly  of  the  unsaturated  class,  and  show  an  affinity 
for  basic  stains,  as  in  the  case  of  the  Nissl  bodies  of  the  nerve-cells, 
and  perhaps  also  the  cytoplasm  of  plasma  cells.  The  great  question 
concerning  the  cytoplasm  is  its  structure — whether  homogeneous, 
alveolar,  areolar,  fibrillar,  foam-like,  or  granular.  On  a  previous 
page  have  been  mentioned  the  experiments  of  Hardy,  which  show 
that  homogeneous  solutions  of  protein,  when  fixed  by  the  same 
reagents  as  are  used  in  the  customary  fixation  of  histological  mate- 
rials, may  show  quite  the  same  microscopical  structures  as  are  shown 
by  the  cytoplasm  of  cells.  Network,  foam,  and  alveolar  structures  are 
produced  in  albumin  and  gelatin  solutions  when  they  are  hardened  by 
bichloride  of  mercury,  osmic  acid,  formalin,  etc.,  and  the  same  char- 
acteristic differences  that  are  produced  in  cells  by  these  different 
reagents  are  likewise  produced  in  the  hai'dened  protein  solution. 
Protein  structures  hardened  under  strain  form  radiating  stmctures 
resembling  centrosomes  and  the  radiating  threads  seen  in  cells.  If 
elder  pith  is  saturated  with  protein  solutions  and  then  hardened,  sec- 
tioned, and  stained  by  the  usual  methods,  aiipearances  resembling 
closely  the  structure  of  a  hardened  cell  may  be  found  in  the  spaces  of 
the  pith — even  a  central,  niu'leus-like  mass  may  be  suspended  in  a 
network  of  anastomosing  threads.     These  and  mnny  other  experiments 

33  Ppntamalli,  .Arfli.  Enlwicko.  u.  Orj:..  1012  CM).  Ill;  :\I((  Iriuloii.  Pror.  Soc. 
Exp.  P.iol.  and  Med..  1010    (7),  111:    llanly.  .Tonr.  riiysiol,,  1013    (47).   lOS. 


HTRUCTUliE  OF  CYTOPLASM  47 

indicate  that  much  of  the  work  done  on  cell  structure  by  means  of 
studies  of  hardened  cells  cannot  be  considered  of  value  in  deciding  the 
structure  of  living  cells;  but,  nevertheless,  the  fact  remains  that  many 
colls  that  can  be  observed  while  alive  and  uninjured  under  tiie 
microscope  are  seen  to  have  a  definite  structure  in  the  cytoplasm,  e.  g., 
sea-urchin  eggs,  which  show  a  characteristic  alveolar  structure. 

A  compromise  view  of  the  structure  of  protoplasm  (and  cytoplasm 
in  particular)  which  takes  account  of  what  appear  to  be  facts  brought 
out  on  both  sides  of  the  question,  is  that  while  in  some  cells  definite 
structural  arrangements  of  the  cytoplasm  exist,  in  most  cells  the 
proteins  are  chiefly  in  a  homogeneous  solution ;  most  of  the  structures 
seen  in  fixed  cells,  except  the  mitochondria,  chromatin  threads,  nuclear 
membrane,  nucleoli,  and  centrosomes,  are  produced  by  the  coagula- 
tion of  the  proteins,  and  are  not  present  during  life.  AYhen  a  frame- 
work does  exist,  it  is  a  fair  inference,  by  analogy  with  the  cell  mem- 
brane and  the  stroma  of  the  red  corpuscles,  that  the  cell  lipoids  are 
largely  responsible  for  its  formation,  and  that  they  form  a  prominent 
part  of  its  composition.  This  question  of  the  presence  or  absence^of 
structure  in  the  cytoplasm  is  of  more  interest  than  as  a  mere  mor- 
phological problem,  for  if  the  cytoplasm  is  subdi"\dded  into  innumer- 
able little  chambers,  each  surrounded  by  a  membrane,  it  is  probable 
that  processes  of  difi'usion  and  conditions  of  osmotic  pressure  will  be 
very  differefct  from  what  they  would  be  if  the  cj'toplasm  wjere  a  simple 
homogeneous  colloid  solution,  like  a  lump  of  semisolid  gelatin  or  agar. 
In  such  colloidal  masses  diffusion  and  osmosis  go  on  almost  as  if  there 
were  no  colloids  in  the  solvent  at  all,  whereas  most  membrane  struc- 
tures that  are  found  in  living  tissues  seem  to  have  a  decidedly  semi- 
permeable character. 

From  what  we  know  at  the  present  time  of  intracellular  physics 
and  chemistry  there  is  no  necessity  for  assuming  that  semipermeable 
septa  exist  within  the  cell.  ■  All  tlie  intracellular  processes  with  which 
we  are  familiar  could  go  on  without  such  structures.  It  is  not  neces- 
sary to  assume  a  compartment  structure  to  explain  the  possibility  of 
different  chemical  reactions  going  on  in  different  parts  of  the  cell  at 
the  same  time,  for  most  of  the  cell  reactions  seem  to  depend  on 
enzymes,  which  we  know  are  not  readily  diffusible  in  solutions  of  col- 
loids, and,  therefore,  might  remain  fi:xed  without  requiring  any  en- 
closing walls  or  retaining  framework.  Certainly,  many  cells  are  free 
from  structural  cytoplasm,  for  we  see  particles  of  solid  matter  moving 
about  within  tliem  quite  freely.  In  some  cells  the  nuclei  migrate 
about  in  the  cell,  as  also  do  digestive  and  excretoiy  vacuoles,  which 
motion  would  seem  to  be  rather  destructive  if  the  protoplasm  had  a 
structure  at  all  permanent. 

When  a  portion  of  the  cytoplasm  is  cut  free  from  the  body  of 
certain  cells  it  at  once  forms  a  round  drop,  just  as  any  insoluble 
fluid  would  do  in  another  of  different  surface  tension,  and  not  at  all 


48  THE    CHEMISTRY    AND    PHYSICS    OF    THE    CELL 

as  if  it  were  Ixtuiid  into  a  fixed  strueture  by  a  framework.  Other 
cells,  however,  retain  their  form  under  tlie  same  conditions.  The 
structure  of  even  so  evidently  complicated  a  cytoplasm  as  that  of 
striated  muscle  fibers  is  in  doubt ;  a  classical  observation  on  this  point 
is  the  passage  of  a  minute  worm  through  the  substance  of  a  muscle- 
cell,  its  progress  being  as  unimpeded  as  if  there  were  no  such  things 
as  disks,  bands,  rods,  and  stria^  in  the  cell.  INIany  features  of 
ameboid  movement  also  seem  to  indicate  that  the  cytoplasm  follows 
much  the  same  laws  as  a  drop  of  fluid  in  a  heterogeneous  medium,  for 
we  can  make  a  drop  of  mercury  or  of  chloroform  in  water,  or  of  oil 
in  weak  alcohol,  react  to  various  stimuli  in  much  the  same  way  that 
an  ameba  would.  If  we  look  upon  the  cytoplasm  as  a  drop  of  emulsion 
colloid,  the  surfaces  of  the  particles  in  the  emulsion  furnish  of  them- 
selves adequate  explanation  of  many  of  the  phenomena  of  isolation 
of  chemical  reactions,  etc.,  without  lacking  in  harmony  with  the  evi- 
dences of  structural  homogeneity.  This  hypothesis  fits  all  sides  of 
the  problem  and  has  many  supporters  at  the  present  time.^* 

The  question  of  structure  in  the  nucleus  is  quite  a  different  matter, 
in  so  far  as  the  chromatin  threads  and  the  nucleolus  are  concerned.  In 
ameboid  movement  the  nucleus  seems  to  play  a  passive  role  and  to 
be  dragged  about  by  the  cytoplasm,  indicating  quite  a  high  degree  of 
rigidity.  It  is  probable,  however,  that  the  achromatic  portion  between 
the  chromatin  threads  and  granules  has  much  the  same  structure  or 
lack  of  structure  as  the  cytoplasm. 

The  inorganic  salts  seem  to  be,  at  least  in  part,  contained  in  the 
cells  in  chemical  combination  rather  than  in  simple  solution  in  the 
water  of  the  cell.  There  is  much  evidence  indicating  that  they  form 
with  the  proteins  ion  compounds,  which  may  be  altered  under  various 
conditions.  For  example,  Loeb  found  that  muscles  placed  in  solutions 
of  potassium  salts  took  up  much  water,  whereas  if  placed  in  a  solution 
of  calcium  salts  they  lost  water,  exactly  as  soaps  do  w^hen  potassium  or 
calcium  ions  are  substituted  for  the  sodium  ions  in  a  sodium  soap. 
He  has  suggest(Ml  that  we  have  in  the  cells  a  protein-ion  compound, 
after  this  order,  Xa 

/ 

Protein — K 

\ 

Ca 

aiid  that  if,  in  the  surrounding  fluid,  a  great  excess  of  one  of  these 
ions  is  present,  it  may  displace  the  others  by  mass  action,  forming  a 
protein  with  all  or  most  of  the  ions  of  one  kind,  and,  therefore,  de- 
cidedly abiioi-mal.  ^Fany  features  of  cell  physiology  seem  explainable 
on  these  o-rouiids,  and  the  reader  is  referred  to  Loeb's  collected  wcu-ks 
for  fnrtlici-   discnssion ;  ^^  also  to   Clowes'  interesting  investigations 

34  An   oxcollont  discussion  of  tliis  (|iu'slinn    is   j^iven   liv    Alsljorji:,   Scioiico.    lOll 
(34),  07. 
30  "Studios  in  Coneral   Pli\>ioi()>'V,"   ]!)05. 


THE  CELL   WALL  49 

on  tlic  influt'iice  of  different  ions  on  emnlsion  formation  and  mem- 
brane l)ermeabilit3^•'''"  The  influence  of  inorganic  salts  on  the  swell- 
ing of  tissue  colloids  is  also  tliscussed  from  another  standpoint  by  M. 
H.  Fischer  in  his  work  on  ''Edema."  In  any  event  it  is  important 
ior  the  cell  that  the  proportion  of  the  inorganic  constituents  be  main- 
tained in  rather  constant  conditions  of  quality  and  ([uantity. 

The  various  secretory  granules,  fat-droplets,  pigment-granules, 
gh'cogen  granules,  keratin,  etc.,  that  may  lie  in  the  cytopla.sm,  are 
inconstant  constituents,  varying  with  different  cells,  and  under  varj^- 
iug  conditions  in  the  same  cells,  and  lie  beyond  the  scope  of  our  dis- 
cussion of  the  general  coin])osition  of  the  cell.  According  to  Ruzicka  ^"^ 
there  is  contained  in  all  cells,  both  in  nucleus  and  cytoplasm,  an  in- 
soluble substance  which  corresponds  structurally  to  the  "plastin"  of 
the  cytologists,  and  chemically  is  related  to  the  reticulins  and  other 
albuminoids;  this  he  looks  upon  as  the  ground  substance  of  the  cells, 
corresponding  to  the  albuminoid  ground  substance  of  stroma  of  or- 
ganized tissues. 

Certain  of  the  granulations  observed  in  the  cj'toplasm  of  cells  seem 
to  be  definite,  constant  structures  of  the  living  protoplasm,  and  these 
are  now  called  mitochondria,  which  term  includes  many  forms  of 
granules  described  under  various  names.^""  Their  solubility  and 
staining  reactions  suggest  that  they  contain  phospholipins,  perhaps 
associated  with  proteins.  Their  functional  importance  is  indicated 
by  the  fact  that  usually  their  number  varies  directly  with  the  meta- 
bolic activity  of  the  cells,  and  they  may  be  related  to  histogenesis. 

Other  histological  cellular  structures  also  permit  of  more  or  less 
satisfactory  identification  by  microchemical  methods,  and  Unna^^*" 
especially  has  contributed  to  this  field.  By  staining  sections  with 
dyes  of  varying  reaction,  after  extracting  the  sections  with  various 
solvents,  he  has  obtained  evidence  of  the  chemical  nature  of  some  of 
the  cell  structures,  although  it  is  by  no  means  certain  that  the  con- 
clusions drawn  will  all  be  verified.  In  the  nucleolus  he  finds  a  sub- 
stance resembling  globulin,  the  granuloplasm  of  the  cell  body  he  re- 
gards as  an  albumose,  the  spongioplasm  as  histone,  mast  cell  granules 
as  mucin  or  mucoid  substances.  Nissl  bodies  he  holds  to  be  albumose, 
altho  others  have  believed  them  to  be  nucleins.^^'^ 

THE   CELL-WALL  37 

The  cell  membrane  in  most  animal  cells  is  inconspicuous  struc- 
turally, but  in  discussing  osmosis  it  was  sho^^^l  that  it  is  of  the  greatest 

35a  Jour.  Physical  Chem.,   1916    (20),  407. 
scArch.  f.  Zellforsch.,  1008   (1),  587. 

36a  See  review  bv  Cowdrv.  Amer.  Jour.  Anat.,  1916   (19),  42.3. 
sfibSee  review  bv  Gans,  "Dent.  med.  Wooh.,  191.3    (39),  1944. 
36c  See  Unna,  Berl.  klin.  Woeh..  1914    (51),  444:  :Miihliiiann.  Arch.  mikr.  Anat.. 
1914    (85).  361. 

3"  See  Zangger,  '"Ueber  ^lembranen  und  IMcjnljrancnfunktionen,"   Krgcbiiisse  d. 
4 


50  THE  CHEMISTRY  AND  PHYSICS  OF  THE  CELL 

biological  importance.  There  is  no  direct  chemical  or  microscopical 
evidence  at  hand  showing  the  composition  of  the  animal  cell  mem- 
brane, but  b}^  observations  on  its  behavior  when  the  cells  are  in  solu- 
tions of  different  sorts,  facts  have  been  collected  indicating  that 
lecitliin  and  cholesterol,  and  probably  the  allied  fat-like  bodies, 
"protagon"  and  cerebrin,  are  prominent  constituents.  The  sub- 
stances that  diffuse  through  most  cell  walls  are  just  the  substances  that 
are  soluble  in  or  dissolve  these  lipoids,  e.  g.,  alcohol,  chloroform,  ether^ 
etc.,  and  it  is  prol)able  that  the  anesthetic  effects  of  many  of  these 
substances  depend  in  some  w'ay  on  their  fat-dissolving  power  and 
the  large  proportion  of  lipoids  in  nerve-cells.  These  observations  were 
made  first  by  Overton  ^^  and  Meyer,^^  and  led  to  the  now  prominent 
but  disputed  hypothesis  that  the  permeability  of  cells  is  determined 
by  the  lipoids.  Of  particular  interest  for  our  purpose  are  Over- 
ton's observations  on  the  effects  of  dyes  on  living  cells.  The  best 
known  vital  stains  {i.  e.,  stains  that  will  enter  the  living  cell  without 
requiring  or  causing  injury  to  it)  are  neutral  red,  methylene  blue,, 
toluidin  blue,  thionin,  and  safranin.  If  uninjured  cells,  e.  g.,  frog 
eggs,  are  placed  in  watery  solutions  of  these  dyes  they  soon  become 
filled  with  the  coloring-matter,  which  seems  to  penetrate  the  cell  mem- 
brane quite  uniformly  at  all  points ;  if  the  dyed  eggs  are  then  placed 
in  clear  water,  the  stain  diffuses  out  again,  show'ing  it  to  be  simply 
absorbed,  rather  than  chemically  combined.  In  contrast  to  these 
stains  the  sulphonic  acid  dyes,  such  as  indigo  carmine  and  water- 
soluble  indulin,  nigrosin,  and  anilin  blue,  do  not  penetrate  the  living 
cell  at  all.  Overton  tested  the  solubility  of  dyes  which  are  not  vital 
stains  and  found  them  all  insoluble  in  oils,  fats,  and  fatty  acids ;  but 
the  dyes  staining  living  cells  were  readily  soluble  in  lecithin,  choles- 
terol, "protagon,"  and  cerebrin,  the  so-called  cell  lipoids.  Further- 
more, if  crumbs  of  lecithin,  "protagon,"  or  cerebrin  were  placed  in 
very  dilute  watery  solutions  of  these  dyes,  they  were  found  to  absorb 
from  the  water  the  vital  stains,  but  not  the  others,  whieli  indicates- 
that  stains  that  penetrate  living  cells  are  more  soluble  in  lipoids 
than  they  are  in  water. 

Many  exceptions  to  this  rule  of  the  fat  solubility  of  dyes  which 
can  penetrate  living  cells  have  been  found,  especially  by  Ruhland,^" 
and  the  universal  applicability  of  the  Overton-IMeyer  hypothesis  has 
been  questioned.  It  is  at  once  evident  that  the  common  foodstuffs 
which  enter  the  cell,  such  as  water,  sugar,  amino-acids,  and  salts  are 
not  li})()id-soluble,  hence  it  has  been  suggested  that  the  cell  membranes 
must  have  a  "mosaic"  striiclui-e,  some  of  the  blocks  being  lipoids  or 

Physiol..  1008  (7),  99;  also  T{.  S.  l.illic.  "TIh'  TJule  of  Moiiibraiios  in  Coll  Proc- 
esses," Popular  Science  Montlily,  Feb.,   I'M:!. 

38.Tahrb.    f.   wissontsohafll.    I'joianik,    1!H)0    (34),    609. 

30  .Arch.  f.  exp.  Patli.  u.  PliarTii.,   1S99    (42),  109. 

4n.Talirb.   f.  Wis.seiischaft.   Botanik,   1912    (f)!),  .-{TO. 


THE  CELL  WALL  51 

lipoid  conipoiinds,  and  others  proteins  without  lipoids,  (Robertson  *^ 
suggests  that  there  is  a  supei-fieial  film  of  concentrated  protein  about 
the  cells,  underlaid  by  a  discontinuous  lipoid  layer.)  There  is,  fur- 
thermore, evidence  that  the  entire  cell  substance  has  a  profound  effect 
upon  diffusion  within  the  cell,  so  that  it  is  at  present  impossible  to 
say  whether  the  osmotic  phenomena  of  cells  depend  upon  a  cell  mem- 
brane or  upon  the  entire  cell  substance. ^^-^  It  may  be  that  there  are 
membranes  or  surfaces  within  the  cell,  as  postulated  in  the  foam 
structure  hypothesis  of  protoplasm,  or  that  a  homogeneous  protoplasm 
develops  surfaces  where  in  contact  with  substances  entering  from  the 
outside. 

Many  facts  indicate  that  either  the  delicate  external  membrane 
of  animal  cells  or  the  entire  cytoplasm  has  the  features  of  a  semi- 
permeable membrane,  to  the  extent  of  permitting  certain  substances 
to  diffuse  through  and  not  others.  Had  they  the  property  of  some 
of  the  artificial  semipermeable  membranes,  of  letting  water  pass 
through  but  holding  back  almost  absolutely  all  crystalloids,  the  re- 
sult would  be  the  development  of  an  enormous  disproportion  in  the 
pressure  between  the  inside  and  the  outside  of  the  cell.  Furthermore, 
the  exchange  of  nutritive  material  and  excretion  products  between  the 
blood  and  the  cells  would  be  impossible.  But  permitting  some  sub- 
stances to  pass  into  the  cell  results  in  their  accumulation  within  the 
cell,  until  they  are  in  sufficient  concentration  to  neutralize  the  osmotic 
pressure  exerted  on  the  outside  of  the  cell.  As  evidence  of  this  elec- 
tive permeability  we  have  the  fact  that  the  proportion  of  certain  salts 
within  the  cell  is  quite  different  from  what  it  is  in  the  fluids  bathing 
them;  e.  g.,  animal  cells  generally  contain  more  potassium  and  less 
sodium  than  the  fluids  surrounding  them.  The  inorganic  constituents 
of  red  cells  are  different  from  those  of  the  plasma,  the  corpuscles 
not  containing  any  calcium  at  all,  while  the  magnesium  seems  to 
enter  them  freely;  in  other  words,  the  red  coi'puscle  seems  to  be 
impermeable  to  calcium  and  permeable  to  magnesium.  If  the  salts 
in  a  corpuscle  are  in  smaller  proportion  than  in  the  surrounding  fluid, 
it  indicates  that  the  cell  membrane  is  not  freely  penneable  for  them ; 
if  in  greater  proportion,  that  some  constituent  of  the  cell  is  holding 
them  in  combination,  possibly  as  ion-protein  compounds.  Probably 
inorganic  salts  are  present  in  the  cell  by  virtue  of  both  physical  and 
chemical  influences,  some  simply  diffusing  in  and  out,  others  com- 
bining with  the  proteins  and  being  held  chemically. 

The  intercellular  substance  varies  greatly  in  different  tissues.  In 
the  case  of  the  supportive  tissues  it  is  the  important  element,  and 
the  cells  seem  to  exist  chiefly  for  the  purpose  of  forming  and  keeping 
it  in  repair  as  it  is  worn  out.  In  the  epithelial  and  secreting  tissues, 
however,  the  intercellular  substance  is  reduced  to  a  minimum,  except 

"Jour.  Biol.  Chem.,   1908    (4),   1. 

4ia  See  Kite,  Amer.  Jour.  Pliysiol.,  1915    (37),  282. 


52  THE  CHEMISTRY  A.AD  PHYSICS  OF  THE  CELL 

in  SO  far  as  a  cement  substance  is  required,  and  the  cells  generally  lie 
in  almost  immediate  apposition.  It  is  probable  that  there  is  a  greater 
or  less  amount  of  cement  substance,  even  between  the  most  closely 
applied  cells,  and  this  substance  seems  to  be  related  to  mucin.  It 
(;an  generally  be  brought  out  by  staining  with  silver  nitrate,  and 
]\racallum  ^"  points  out  that  this  reaction  is  merely  a  micro-chemical 
test  for  chlorides,  and  indicates  that  the  cement  substance  contains 
them  in  larger  proportion  than  does  the  cytoplasm. 

42  Proceedings  of  the  Royal  Society,  1905  (76),  217. 


CHAPTER   II 

ENZYMES 

Every  cell  is  eoiistaiitly  aeeoinplishin<>:  an  enormous  luiinber  of 
chemical  reactions  of  varied  natures,  at  one  and  the  same  time ;  how 
many  we  do  not  know,  but  the  score  or  more  that  we  do  know  to  be 
constantly  going  on  in  the  liver  cell,  for  example,  are  probably  but 
a  part  of  the  whole.  Furthermore,  reactions  take  place  between  sub- 
stances that  show  no  inclination  to  affect  each  other  outside  the  body, 
and  proceed  in  directions  that  we  find  it  difficult  to  make  them  take 
in  the  laboratory.  Proteins  are  being  continually  broken  down  into 
urea,  carbon  dioxide,  and  water ;  yet  to  split  proteins  even  as  far  as  the 
amino-acid  stage  requires  prolonged  action  of  concentrated  acids  or 
alkalies,  or  super-heated  steam  under  great  pressure.  But  all  the  time 
in  the  cell  a  multitude  of  equally  difficult  changes  is  going  on  at  once, 
within  its  tiny  mass,  always  keeping  the  resulting  heat  within  a  frac- 
tion of  a  degree  of  constant,  and  the  resulting  products  within  narrow 
limits  of  concentration.  We  have  already  indicated  the  means  used 
to  keep  the  concentration  of  the  cell  products  withm  safe  limits; 
namely,  the  processes  of  diffusion  and  osmosis  and  their  modification 
by  the  cell  structure.  The  forces  that  bring  about  the  chemical  reac- 
tions reside,  we  say,  in  enzymes,  although  in  so  doing  we  only  shift 
the  attribute  formerly  conceded  to  the  cell,  to  certain  constituents  of 
the  cell  whose  nature  and  manner  of  action  are  equally  unkno\\ai. 
AVlien  the  only  enzymes  that  were  known  were  limited  to  those  se- 
creted from  the  cell,  and  found  free  in  fluids,  such  as  pepsin  and  tryp- 
sin, the  chemical  changes  that  went  on  in  the  cell  were  ascribed  to  its 
"vital  activity."  Buchner,  by  devising  a  method  to  crush  j^east  cells, 
and  finding  the  expressed  cell  contents  able  to  produce  the  same 
changes  in  carbohydrates  that  the  cells  themselves  did,  ]n'oved  the  ex- 
istence within  living  cells  of  enzymes  similar  to  those  excreted  by  cer- 
tain cells,  and  substantiated  the  belief  of  their  existence  that  had 
become  general  before  it  was  thus  finally  corroborated.  Growing  out 
from  this  and  subsequent  experiments  has  come  a  larger  and  larger 
amount  of  evidence  that  many  of  the  chemical  activities  of  the  cells 
are  due  to  the  enzymes  they  contain,  until  now  the  point  is  reached 
where  one  may  rightfully  ask  if  cell  life  is  not  entirely  a  matter  of 
enzyme  activity.  There  are  certain  facts,  however,  which  seem  to  in- 
dicate that  there  are  some  essential  differences  between  cells  and 
enzymes.     One  of  the  most  importaiit  of  these  is  the  difference  in  the 

5.3 


54  ENZYMES 

susceptibility  to  poisons  of  enzymes  and  cells.^  Strengths  of  certain 
antiseptics  that  will  either  destroj^  or  inhibit  the  action  of  living  cells, 
such  as  alcohol,  ether,  salicylic  acid,  thymol,  chloroform,  toluene  and 
sodium  fluoride,  will  harm  free  enzymes  in  solution  little  or  not  at  all. 
This  fact  has  been  of  great  assistance  in  distinguishing  between  the 
action  of  enzymes  and  of  possible  contaminating  bacteria  in  experi- 
mental work.  Although  this  difference  between  enzymes  and  cells  is 
characteristic,  it  does  not  finally  decide  that  tlie  cell  actions  are  not 
enzyme  actions,  for  it  may  well  be  that  the  poisons  act  chiefly  by 
altering  the  physical  conditions  of  the  cell  so  that  diffusion  is  inter- 
fered with,  thus  seriously  interfering  with  the  exchange  of  cleavage 
products  between  different  parts  of  the  cell,  and  cheeking  intracellu- 
lar enzyme  action,  which  we  shall  see  later  requires  free  diffusion  of 
the  products  for  its  continuance.  At  the  very  least,  however,  we  may 
look  upon  the  intracellular  enzymes  as  the  most  important  known 
agents  of  cell  metabolism,  and  consequently  of  all  life  manifestations, 
and  the  changes  they  undergo  or  produce  in  pathological  conditions 
must  be  fully  as  fundamentally  important  as  is  their  relation  to 
physiological  processes.  It  therefore  becomes  necessary  for  us  to 
consider  carefully — 

THE  NATURE  OF  ENZYMES  AND  THEIR  ACTIONS  - 

Since  up  to  the  present  time  no  ferment  has  been  isolated  in  an 
absolutely  pure  condition  we  are  entirely  unfamiliar  with  their  chemi- 
cal characters,  and  consequently  are  obliged  to  recognize  them  solely 
by  their  action.  As  far  as  we  know,  true  enzymes  never  occur  except 
as  the  result  of  cell  life — they  are  produced  within  the  cell,  and  in- 
creased in  aniount  by  each  new  cell  that  is  formed,  and,  furthermore, 
they  are  present  in  every  living  cell  without  exception.  As  the  same 
facts  are  equally  true  of  the  proteins,  and  apjiarently  true  of  nothing 
else,  it  is  natural  to  associate  the  enzymes  with  proteins,  and  so  ex- 
plain the  importance  of  the  proteins  for  cell  life.^  If  enzymes  are 
obtained  in  any  of  the  usual  ways  from  animal  cells  or  secretions 
they  are  always  found  to  give  the  reactions  for  proteins,  even  if  re- 
purified  many  times.  But  it  is  well  known  that  whenever  proteins 
are  precipitated  the  other  substances  in  the  solution  tend  to  bo  dragged 

1  See  discussion  by  Vernon,  Er<?Gbniase  d.  Pliysiol.,  1910   (0),  2.34. 

2  It  would  not  bo  profitable  to  discuss  fnllv  all  the  various  tlioorics  and  hy- 
potheses that  have  been  advanced,  but  the  reader  is  referred  to  the  followinir  chief 
compilations  of  tlie  entire  subject:  Oppcnlieinier,  '"Die  F(M-nienlc  uiid  ihre  \Mrk- 
unpen,"  Leipzip;  P>ayliss,  "The  Nature  of  l^'n/.ynie  AcUon."  I\l(Uioij;ra|)lis  on  Bio- 
chemistry, London:  Stern,  "Pliysico-cliemical  Pasis  of  I'erinent  Action,"  in  Oppen- 
heimer's  "Ilandbuch  d.  P.iochemie,"  Vol.  4,  pt.  2:  Samuelcy,  "Animal  Ferments." 
ibid.  Vol.  I;  A.  E.  Taylor,  "On  Fermentation,"  T^niv.  of  California  Publications; 
Euler,  "General  Chemistrv  of  tlie  Enzvmes,"  translated  bv  T.  IT.  Pope,  New  York, 
1912. 

3  Another  impfirtaiit  point  is  that  the  closest  imitation  of  cn/yin(>s,  Bredig's 
"inorganic  ferments,"  seem  to  owe  their  action  to  th(>ir  colloidal  nature. 


PROPERTIES  OF  EX/A.UES  55 

down  by  the  colloids,  and  it  is  possible  that  the  enzymes  are  merely 
associated  with  the  proteins  in  this  way.  Furthermore,  enzymes  are 
known  to  become  so  closely  attached  to  stringy  protein  masses,  such  as 
fibrin  and  silk,  that  they  cannot  be  removed  by  washing.  Some  have 
claimed  that  they  have  secured  active  preparations  of  pepsin  and 
invertase  that  did  not  give  protein  reactions  and  contained  very  little 
or  no  ash  or  carbohydrate ;  but  it  has  so  far  been  impossible  to  secure 
trypsin  free  from  protein,  and  diastase  seems  to  be  certainly  of 
protein  nature.  Analyses  of  enzymes  purified  as  completely  as  pos- 
sible do  not  have  great  worth,  for  the  ''purified"  enzymes  are  prob- 
ably far  from  pure;  however,  it  is  of  some  importance  that  they  vary 
greatly  in  the  proportions  of  carbon,  hydrogen,  and  nitrogen  which 
they  contain,  indicating  that  possibly  different  enzymes  may  be  of 
very  different  nature.  The  enzymes  have  been  found  to  possess  defi- 
nite electrical  charges,  in  neutral  solutions  trypsin  is  negative  or 
amphoteric,  pepsin  and  invertase  negative  (INIichaelis).*  jNIacallum 
has  shown  microchemieally  that  phosphorus  is  closely  associated  with 
the  formation  of  zymogen  granules  in  cells,  which  seem  to  be  started 
in  the  nucleus;  and  there  are  many  other  observations  suggesting 
that  certain  ferments  are  closely  related  to  the  nucleo-proteins.  This 
is  particularly  true  of  the  oxidases,  which  seem  also  to  contain  iron 
and  manganese.  A  final  point  of  importance  in  support  of  the  pro- 
tein nature  of  enzymes  is  that  pepsin  destroys  tr\'psin  and  diastase, 
while  trypsin  destroys  pepsin. 

So  uncertain,  however,  is  our  information  concerning  the  chemical 
nature  of  the  enzymes,  that  it  has  become  possible  for  an  hypothesis  to 
be  developed  urging  that  enzymes  are  immaterial,  that  the  actions 
Ave  consider  as  characterizing  enzj^mes  are  the  result  of  physical  forces 
which  may  reside  in  many  substances,  and  perhaps  even  free  from 
visible  matter,  but  the  weight  of  evidence  at  present  available  is  en- 
lirel}'  in  favor  of  the  view  that  enzjones  are  colloidal  substances,  al- 
though perhaps  of  widely  differing  chemical  nature.  A  valuable  piece 
of  evidence  of  the  material  existence  of  enzymes  is  their  specific  na- 
ture, lipase  affecting  only  fats,  and  trypsin  only  proteins,  indicating 
chemical  individuality.  They  are  true  secretions,  formed  A\ithin  the 
cell  by  recognizable  steps;  and,  furthemiore,  when  injected  into  the 
body  of  an  animal,  they  give  rise  to  the  formation  of  specific  innnune 
bodies  that  antagonize  their  action.  Emil  Fischer's  work  with  the 
sugar-splitting  enzymes,  moreover,  indicates  that  they  owe  their  action 
to  their  stereochemical  configiiration.  He  prepared  two  .sets  of  sugar 
derivatives  which  differed  from  each  other  solelj'  in  the  arrangement 
of  their  atoms  in  space  (i.  e.,  isomers)  and  found  that  one  specific 
enzyme  w^ould  split  members  of  only  one  of  the  varieties,  while  nn- 
other  enzyme  would  act  only  on  the  variety  with  the  opposite  isomeric 
form.  These  experiments  make  it  very  probable  that  there  must  be  a 
^Biochem.  Zeit.,  1909   (16),  81  and  486;    (17),  231. 


56  EXZYMEfi 

certain  relation  of  jreometrieal  stnicture  between  an  enzyme  and  the 
substances  it  acts  upon,  and  leaves  little  question  of  its  material  na- 
ture. 

Bredif?  has  found  that  colloidal  sohifio)is  of  metals  have  many  of 
the  properties  of  true  enzymes,  aeeomi)lishino-  many  of  the  decom- 
positions produced  by  enzymes,  being  affected  by  temperatures  of 
nearly  the  same  degree,  and  even  being  ''poisoned"  by  substances 
that  destroy  or  check  enzymes.  The  only  possible  explanation  of  these 
observations  seems  to  ])e  that  the  enzyme  effects  are  brought  about  by 
surface  phenomena.  A  colloidal  solution  of  platinum,  so  far  as  is 
known,  differs  from  a  piece  of  metallic  platinum  solely  in  the  enor- 
mously great  amount  of  surface  it  offers  in  proportion  to  its  weiglit, 
and  it  is  well  known  that  surfaces  may  affect  chemical  action. 
Hence  we  have  the  possibility  that  some  enzyme  actions,  at  least, 
may  depend  upon  the  existence  of  a  very  large  surface,  and  since 
by  no  means  all  colloids  are  enzymes,  that  this  surface  must  bear  a 
certain  relation  in  form  to  the  surface  of  the  body  that  is  to  be  acted 
upon. 

THE  PRINCIPLES  OF  ENZYME  ACTION 

The  effects  produced  by  enzymes,  which  at  one  time  were  con- 
sidered ciuite  unique  and  remarkable,  have  now  been  made  compara- 
tively plain,  chiefly  through  the  observations  of  Ostwald  on  related 
chemical  reactions ;  and  by  the  investigations  of  Croft  Hill,  Kastle 
and  Loevenhart,  and  others,  on  enzymes,  which  show  that  enzyme 
action  is  in  no  way  different  from  chemical  action  observed  independ- 
ent of  enzymes.  The  fundamental  consideration  is  that  chemical  re- 
actions are  reversible,  that  is,  that  their  tendency  is  to  establish  an 
equilibrium,  and  that  the  change  may  be  from  either  side  of  the  equa- 
tion.^ The  action  of  enzymes  is  similar  to  that  of  all  catalytic  agents, 
that  is,  they  increase  the  speed  of  reaction.  In  the  case  of  such  a 
reaction  as  that  of  NaOH  and  HCl,  the  reaction  is  so  ra])id  that  the 
effect  of  catalyzers  could  hardly  be  noticed ;  but  with  many  other 
substances  the  reaction  is  very  slow,  and  without  the  ])resence  of 
catalyzers  it  would  go  on  almost  or  quite  imperceptibly.  For  ex- 
ample, ethyl  butyrate  saponifies  on  the  addition  of  water  according 
to  the  following  equation  : 

CJI— 0— OC— CJI:   -f-   R,O^CJI,OH  +  IIOOC— C,H;. 

On  the  other  hand,  if  ethyl  alcohol  aiid  butyric  acid,  the  products 
of  this  reaction,  are  placed  together,  they  will  combine  to  form  ethyl 
butyrate;  in  otlier  words,  the  reaction  is  rexci-sible,  as  indicated  by 
the  arrows  in  the  e(|uation.  In  any  event,  however,  the  reaction  is  not 
complete,  but  continues  only  until   a  certain   deHnite   pro})ortion   of 

5  See  Taylor,  Arcli.  Jut.  ^\vd.,  IJIOS   (2),  14S 


THE  Ph'i\rfpLi:s  OF  Kx/.VMi:  AcTioy  57 

ethyl  alcohol,  butyric  acid,  etliyl  l)utyrati',  and  water  exists.  -wIk'h  the 
chaiijro  will  stop,  i.  e.,  eqiiUibriKin  is  cstubiishcd.  The  time  that 
would  be  required  for  this  reaction  to  occur  at  room  temperature 
would  be  extremely  long,  the  change  being  hardly  noticeable,  but  in 
the  presence  of  a  catalytic  agent  the  reaction  goes  on  much  more 
rapidly.  Catalytic  agents,  therefore,  merely  hasten  reactions  which 
would  go  ou  without  them,  and  they  do  not  initiate  or  change  the  na- 
ture of  chemical  reactions  at  all.  AVhen  equilibrium  is  established,  the 
reaction  stops  and  the  en/ymc  has  notliing  more  to  do.  Furthermore, 
and  this  is  a  recently  a])preciated  fact,  enzymes  will  hasten  synthesis 
just  as  well  as  they  hasten  catalysis.  Croft  Hill  first  showed  that 
maltase  would  synthesize  glucose  into  maltose ;  Kastle  and  Loevenhart 
soon  after  established  the  synthesis  of  ethyl  butyrate  under  the  in- 
fluence of  lipase.  Taylor  '^  first  synthesized  one  of  the  normal 
body  fats,  triolein,  by  the  action  of  lipase  (from  the  ca.stor-oil  bean) 
iipon  oleic  acid  and  glycerol.  Successful  synthesis  of  fats  by  pan- 
creatic lipase  is  described  by  Lombroso.'^  It  may  seem  improbable  at 
first  sight  that  the  synthesis  of  proteins  can  be  accomplished  by 
enzymes,  as  is  the  relatively  very  simple  synthesis  of  carbohydrates 
and  fats,  but  the  improbability  disappears  when  we  recall  that  the 
jjroducts  of  protein  cleavage  are  reconverted  into  body  proteins  after 
absorption  from  the  intestines.  Proteins  manifestly  are  synthesized 
and  we  have  not  a  little  reason  to  believe  that  this  is  accomplished  by 
enzymes,  presumably  by  a  reversal  of  their  action  in  the  establishment 
of  equilibrium.  Taylor  ^  was  able  to  synthesize  protamin,  one  of  the 
simplest  proteins,  by  the  action  of  trypsin  upon  its  cleavage  products, 
and  it  has  been  found  that  the  addition  of  proteolytic  enzymes  to  solu- 
tions of  pure  albumose  leads  to  the  formation  of  a  jelly-like,  insoluble 
protein  substance,  ' '  plastein, ' '  which  seems  to  be  the  effect  of  a  reversed 
action  on  the  part  of  the  enzymes."  Another  well  known  synthetic  ac- 
tion that  seems  to  be  due  to  reversible  ferment  action  is  the  formation 
of  hippuric  acid  from  benzoic  acid  and  glycocoU  in  the  kidney;  the 
formation  of  glucose  into  glycogen  and  its  reformation  are  also  prob- 
ably both  accomplished  by  one  and  the  same  enzyme  acting  reversibly. 
Other  reversible  reactions  less  closely  related  to  animal  cells  liave  also 
been  described. 

The  reversible  nature  of  enzyme  action  explains  many  problems  of 
metabolism,  and  makes  the  whole  field  much  clearer.  The  following 
consideration  of  the  newer  understanding  of  fat  metabolism  on  this 

6  Univ.  of  California  Publications    (Pathology),  1004   (1),  33. 

7  Arch,  di  farmacol.,  1012   (14),  420. 
8. Tour.  Biol.  Cliem..  1000    (r,),  381. 

0  See  Micheli,  Arch.  ital.  biol.,  1000  (46).  185:  Lcvcnc  aiul  Van  Slykc.  r.iocliom. 
Zeit,  1008  (13),  4.58;  Tavlor,  .Tour.  Biol.  Cheni..  1000  (5),  .300;  Gay  and  Robert- 
son, ibid.  1912  (12),  233;  Abderhalden.  Fermentforsch.,  1014  (1),  47;  v.  Knaffl- 
Lenz  and  Pick,  Arch.  exp.  Path.,  1013   (71),  206,  407. 


58  ENZYMES 

beisis  may  explain  the  manner  in  which  chemical  changes  are  believed 
to  occur  in  the  cells  and  fluids  of  the  body :  " 

In  the  intestines  fat  is  split  by  lipase  into  a  mixture  of  fat,  fatty  acid,  and 
glj'cerol;  but  as  the  fatty  acid  and  glycerol  are  diffusible,  while  the  fat  is  not, 
they  are  separated  from  the  fat  by  absorption  into  the  wall  of  the  intestine. 
Hence  an  equilibrium  is  not  reached  in  the  intestine,  so  the  splitting  continues 
until  practically  all  tiie  fat  has  been  decomposed  and  the  products  absorbed. 
When  this  mixture  of  fatty  acid  and  glycerol  first  enters  the  epithelial  cells  lining 
the  intestines  there  is  no  equilibrium,  for  there  is  no  fat  absorbed  with  them  as 
such.  Therefore  the  lipase,  which  Kastle  and  Loevenhart  showed  was  present 
in  these  cells,  sets  about  to  establish  equilibrium  bj'  combining  them.  As  a  result 
we  have  in  the  cell  a  mixture  of  fat,  fatty  acid,  and  glycerol,  which  will  attain 
equilibrium  only  when  new  additions  of  the  two  last  substances  cease  to  enter 
the  cell.  Now  another  factor  also  appears,  for  on  the  other  side  of  the  cell  is 
the  tissue  fluid,  containing  relatively  little  fatty  acid  and. glycerol.  Into  this  the 
diffusible  contents  of  the  cell  v.'ill  tend  to  pass  to  establish  an  osmotic  e(iuilibrium, 
which  is  quite  independent  of  the  chemical  equilibrium.  This  abstraction  of  part 
of  the  cell  contents  tends  to  again  overthrow  chemical  equilibrium,  there  now  being 
an  excess  of  fat  in  the  cell.  Of  course,  the  lipase  will,  under  this  condition,  reverse 
its  action  and  split  the  fat  it  has  just  built  into  fatty  acid  and  glycerol.  It  is 
evident  that  these  processes  are  all  going  on  together,  and  that,  as  the  composi- 
tion of  the  contents  of  the  intestines  and  of  tlie  blood-vessels  varies,  the  direction 
of  the  enzyme  action  will  also  vary.  In  the  blood-serum,  and  also  in  the  lym- 
phatic fluid,  there  is  more  lipase,  which  will  unite  part  of  the  fatty  acid  and 
glycerol,  and  by  removing  them  from  the  fluid  abotit  the  cells  favor  osmotic  diffu- 
sion from  the  intestinal  epithelitmi,  thus  facilitating  absorption. 

Quite  similar  must  be  the  process  tliat  takes  place  in  the  tissue  cells  through- 
out the  body.  In  the  blood-serum  batliing  the  cells  is  a  mixture  of  fat  and  its 
constituents,  probably  nearly  in  equilibrium,  since  lipase  accompanies  them.  If  the 
diffusible  substances  enter  a  cell  containing  lipase,  e.  g.,  a  liver  cell,  the  process 
of  building  and  splitting  will  be  quite  the  same  as  in  the  intestinal  epitlielium. 
The  only  difference  is  that  here  the  fatty  acid  may  be  removed  from  the  cell  by 
being  utilized  by  oxidation  or  some  other  chemical  transformation.! i 

To  summarize,  it  may  be  stated  that  throughout  the  body  there  is 
constantly  taking-  place  both  splitting  and  building  of  fat.  Fat  enters 
the  cells,  leaves  them,  and  is  utilized  only  in  the  form  of  its  acid  and 
alcohol,  never  as  the  fat  itself.  Fat  constitutes  a  resting  stage  in  its 
own  metabolism. 

If  proteolytic  enzymes  also  act  reversibly,  then  the  phenomena  of 
protein  metabolism  are  similarly  explained,  for  there  is  no  doubt 
that  every  cell  and  body  fluid  contains  proteolytic  enzymes. 

All  metabolism,  then,  may  be  considered  as  a  continuous  attempt  at 
establishment  of  equilibrium  bij  enzymes,  perpetuated  by  prevention 
of  attainment  of  actual  equilibrium  through  destruction  of  some  of 
the  participating  substances  by  oxidation  or  other  chemical  processes, 

10  See  Loevenhart,  Amer.  Jour,  of  Physiol.,  1002  (G),  331;  Wells,  Journal 
Amer.  Med.  Assoc,  1902  (38),  220.  The  discrepancies  between  tiie  action  of 
lipase  in  the  tissties  and  in  vitro  are  well  explained  liy  Taylor,  Jour.  Biol.  Chem., 
1906    (2),   103. 

11  Bradley  (.Tour.  Biol.  Chem.,  1910  (8),  2r)l;  1913  (13).  407-439)  calls  atten- 
tion to  tlic  great  conc(>ntration  necessary  for  fat  synthesis  by  lipase  in  vitro, 
and  tlie  lack  of  correspondence  between  Die  amount  of  fat  and  of  lipase  in  various 
tissues,  questioning  tlie  importance  of  lipase  for  fat  synthesis  in  the  living  tissues 
as  well  as  the  significance  of  reversed  enzyme  reaction  for  biological  processes  in 
general. 


GENERM.  i'if<)i'i:irrii:s  or  i:\/.y\ii:h  59 

or  by  removal  from  the  cell  or  entrance  into  it  of  materials  which  over- 
halance  one  side  of  the  equation. 

In  just  Avhat  manner  the  enzymes  accomplish  tlicir  catalytic  effect 
is  yet  unknown.^-  A  favorite  idea  is  that  they  form  loose  compounds 
Avith  the  substance  to  be  split  and  with  water ;  the  resulting  compound 
being  unstable  and  breaking  down,  the  water  remains  attached  to 
the  components  of  the  substance. 

Enzymes  do  not  act  eatalytically  on  all  substances  by  any  means, 
but  show  a  decidedly  specific  nature.  The}'  affect  only  organic  sub- 
stances, and  the  actions  are  limited  to  two  processes — hydrolysis  and 
oxidation,  or  the  reverse  processes  of  dehydration  and  reduction.^^ 
The  most  essential  difference  between  the  enzymes  and  the  chemicals 
that  can  accomplish  hydrolysis  or  oxidation  is  this :  the  ordinary 
-chemical  reagents  produce  their  effects  on  many  sorts  of  substances, 
whereas  the  enzymes  are  specific ;  thus  hydrochloric  acid  will  hydrolyze 
starch  or  protein  with  equal  facility,  but  pepsin  will  not  affect  starch 
at  all. 

The  very  specific  nature  of  the  enzymes,  their  activation  by  other 
l^ody  products,  the  fact  that  they  seem  to  be  bound  to  the  substance 
upon  which  they  act,  that  they  are  susceptible  to  heat,  and  that  they 
produce  immune  bodies  when  injected  into  experimental  animals,  all 
suggest  the  probability  of  a  relationship  hettveen  enzymes  and  toxins. 
This  matter  will  be  discussed  more  fully  in  considering  the  chemistry 
of  immunity  against  enzymes. 

General  Properties  of  Enzymes. — Other  properties  of  enzymes 
may  be  briefly  mentioned.  The  speed  of  reaction  they  produce  in- 
creases with  the  amount  of  enzymes  present,  but  not  in  direct  propor- 
tion (except  with  rennin).  Very  dilute  acids  favor  the  action  of 
nearly  all  fennents,  and  alkalies  are  unfavorable  for  all  but  trypsin, 
ptyalin,  and  a  few  others.  Weak  salt  solutions  also  are  more  favor- 
able than  distilled  water.  (These  facts  suggest  strongly  the  possi- 
l)ility  that  ions  play  an  important  role  in  the  process.)  Water  and 
dilute  glycerol  dissolve  enzymes,  which  form  always  colloidal  solutions 
that  are  very  slightly  dialyzable ;  and  they  may  be  precipitated  from 
solution  by  alcohol,  and  redissolved  again  with  but  slight  impairment 
of  strength.  Filtration  through  porcelain  filters  is  not  complete,  from 
10  to  25  per  cent,  of  most  enzymes  being  lost  in  each  filtration  and 
enzj'mes  are  subject  to  great  absorption  by  surfaces,  e.  g.,  charcoal, 
kaolin."  As  before  mentioned,  many  chemicals  poisonous  to  bacteria 
have  little  influence  on  most  enzymes,  but  nearly  all  substances  when 
concentrated   are   injurious   or   destructive,    and   some   enzymes   are 

12  See  Euler,  "Chemical  Dynamics  of  Enzyme  Reactions."  Ergebnisse  d.  Pliysiol., 
1910    (9),  241.  *  "  .  _ 

13  Alcoholic  fermentation  may  be  an  exception,  the  change  being  C„II,;Oc  — 
2CoH,,  -f  2C0.,,  but  it  is  very  possibly  an  intramolecular  oxidation. 

14  See  Hedin,  Ergebnisse  d.  Phvsiol'.,  1910   (9),  433. 


60  ENZYMES 

known  that  are  more  susceptible  to  antiseptics  than  are  the  cells  that 
contain  tliein.  Formaldehyde  is  very  destructive  to  enzymes,  even 
when  dilute.  The  efifect  of  protein-coag-ulatiu^  antiseptics  upon  en- 
zymes is,  of  course,  greatly  modified  by  tlie  amount  of  protein  sub- 
stances mingled  with  the  enzymes;  and  the  effects  of  heat  and  other 
injurious  influences  are  greatly  decreased  by  the  presence  of  proteins 
and  other  impurities. 

All  enzymes  are  most  active  between  35°  and  45°  C,  and  it  is  inter- 
esting to  note  that  Kobert  found  this  equally  true  for  enzymes  derived 
from  cold-blooded.animals.  Although  enzymes  can  stand  temperatures 
of  100°' C.  or  more  when  dry,  in  water  they  are  generally  destroyed 
somewhat  below  70°  C.  Low  temperature,  even  —  190° C,  (liquid 
air),  does  not  destroy  them.  The  loss  of  power  through  heating 
occurs  gradually,  and  there  is  no  sharp  line  at  which  their  action 
disappears.  Sunlight  is  harmful  to  enzymes  in  solution,  but  only  in 
the  presence  of  oxygen ;  this  effect  is  augmented  by  the  presence  of 
fluorescent  substances.  Nascent  oxygen  is  destructive  to  enzymes.^** 
Radium  and  a:;-rays  seem  to  have  a  deleterious  effect  upon  most  en- 
zymes, and  retard  their  rate  of  action;  but  apparently,  autolytic  en- 
zymes (Neuberg^")  and  tyrosinase  (Willcock^")  are  not  injured  by 
these  agencies.^*"*  Ultra  violet  rays  are  also  injurious  to  enzymes,^^ 
and  they  can  be  destroyed  by  violent  shaking  (Shaklee  and  INIeltzer  ^^). 
Labile  as  enzymes  are,  their  persistence  when  diy  is  remarkable ; 
Kobert  found  active  trypsin  in  the  bodies  of  spiders  that  had  been  in 
the  Nuremberg  ]\Iuseum  for  150  years,  and  Sehrt  ^^  found  that  the 
muscle  tissue  of  mummies  contained  active  glycolytic  ferment. 

All  enzymes  as  ordinarily  prepared  have  the  property  of  decom- 
posing hydrogen  peroxide,  a  property  possessed  by  substances  of 
varied  nature ;  this  effect  is  prevented  by  CNIT,  wOiich  does  not  pre- 
vent other  enzyme  manifestations,  indicating  that  this  property  is  due 
to  an  associated  enzyme,  catalase. 

The  retardation  of  enzyme  action  by  aecuinuhitioii  oi"  the  products 
of  their  action  is  simply  explained  as  being  due  to  establislnnent  of 
o((uilibrium ;  in  some  instancies,  however,  tlie  substances  jiroduced  are 
of  themselves  harmful  to  tlie  enzymes,  c.  r/.,  alcohol  and  acetic  acid. 

Activation  of  Enzymes. — AVithin  the  cell,  the  enzymes — at  least  those 
1hat  are  excreted,  sncli  as  trypsin  and  pejisin — exist  with  few  excep- 
tions ill  ;iii  active  form,  the  z]iu\()()cu.  Tlieir  activation  a|)pears  to 
take  i)lace  normally  only  after  they  have  been  discharged  from  the 

34a  See  Buifje.  Anicr.  Jour.  I'livsiol,  1014    (34).  140. 

i--Ecr].  klin.  Wocli.,  in()4    (41),   lOSl. 

in  Jour,  of  Pliysiol.,   lOOG    (.34),  207. 

i'!ar;ii(lz('ii1  (Zcil.  Siriililoiitlicr..  1914  (4).  (KKI)  denies  lliiil  rndium  nets  on 
ciizyiiies. 

1"  Ajriillidii.  Ann.  Inst,  rusteur,  lill-J  (1^(1).  .'iS ;  Hiir^'e  i  /  <(/.,  Anier.  .lour. 
PliVBiol..    l!)lt)    (40),  42(i. 

i"s  Ainer.  .T<mr.  Plivsiol.,   1!I0!1    (25),  SI. 

1^'  Meil.   klin.   Wdcii..    I'.MII    (111,    11)7. 


TOXICITY  (IF  l:\y.V MES  61 

cell,  but  after  the  death  of  an  or^aii  it  may  result  from  tin;  decom- 
position products  tliat  are  formed.  I'lider  i)hysiolo<rical  conditions 
this  activation  appears  to  be  brouj^ht  about  by  special  activating,'  sub- 
stances. In  the  case  of  tlie  pancreas  it  is  the  enterokinase,  which  is 
furnished  by  tlie  epithelial  cells  of  the  intestine.  Enterokinase  ap- 
pears to  unite  with  trypsinogen  to  fonn  an  active  enzyme,  which  re- 
minds one  of  the  way  that  complement  and  the  intermediary  l)ody 
unite  to  form  hemolytic  and  bacteriolytic  substances.-"  Kinases,  hav- 
ing the  same  action  as  enterokinase  upon  the  trypsinogen,  are  found 
in  various  tissues  and  organs,  but  generally  much  less  active  than  the 
enterokinase.  Pepsinogen  is  probably  activated  by  the  IICl  of  the 
gastric  juice.  It  is  very  probable  that  it  is  through  this  mechanism 
that  the  rate  of  enzyme  action  is  modified,  and  perhaps  it  is  a  means 
of  defense  of  the  body  against  its  own  enzymes;  as  the  prozymes  are 
more  resistant  to  harmful  agencies  than  the  enzymes,  it  also  may  be 
a  method  of  storage.  The  activity  of  various  enzymes  is  greatly  in- 
creased by  certain  more  or  less  specific  substances,  referred  to  usually 
as  "co-enzymes";  thus  bile-salts  act  as  co-enzymes  for  lipase 
(Loevenhart). 

THE  TOXICITY  OF  ENZYMES 

Although  present  normally  in  greater  or  less  amounts  in  all  the 
cells  in  the  body,  when  artificially  isolated  and  injected  directly  into 
animals  nearly  all  enzymes  seem  to  be  extremely  toxic.  As  foreign 
proteins,  especially  extracts  of  tissues,  are  generally  more  or  less 
toxic,  it  is  difficult  to  state  how  much  of  the  toxicity  of  a  given  enzyme- 
containing  solution  depends  on  the  enzyme  and  how  much  on  the 
admixt  proteins.  The  following  statements  are  taken  at  the  face 
value  placed  on  them  by  the  several  investigators  quoted,  and  are 
subject  to  discount  until  the  enzymes  have  been  isolated  and  investi- 
gated in  a  pure  condition,  if  such  a  thing  shall  ever  become  possible. 

The  first  thorough  study  of  the  toxicity  of  enzymes  was  made  by 
Hildebrandt,-^  who  found  that  pepsin,  invertase,  diastase,  emulsin, 
myrosin,  and  rennin  were  all  toxic.  The  symptoms  produced  in  dogs 
were  trembling,  uneasiness,  difficulty  in  walking,  and  finally  coma. 
The  anatomical  changes  observed  w^ere :  numerous  hemorrhages 
throughout  the  body,  fatty  degeneration  of  the  liver  and  myocardium, 
renal  congestion,  and  numerous  thromboses.  Considerable  fever  re- 
sults, and  ]Mayer  considers  this  responsible  for  the  relative  harmless- 
ness  of  rennin,  the  action  of  which  is  impaired  above  40°,  That  these 
effects  are  due  to  the  enzymes  themselves  rather  than  to  contaminating 

20  Bayliss  and  Starlinp:  (.Tour,  of  Physiol..  Ifl05  (32).  120),  quoslioii  tlu> 
analogy  of  zymogen-kinase  combinations  to  coniplement-amboceptor  eomliination. 
Walker,  however,  finds  evidence  that  many  enzymes  consist  of  a  specitic  ambo- 
ceptor and  a  non-specific  complement  or  kinase  (Jour,  of  Physiol..  IKiKl  (.3.3), 
p.  xxi.). 

2iVircho\v's  Archiv,  1800    (121),   1. 


62  ENZYMES 

bacteria  is  shown  by  Kionka  and  by  Achalme  --  who  obtained  similar 
results  witli  enzymes  made  sterile  by  filtration  through  porcelain. 
Achalme  found  that  such  sterile  preparations  of  pancreatic  juice  in- 
jected subcutaneously  into  guinea-pigs  produce  a  marked  local  pink 
gelatinous  edema,  followed  by  gangrene;  if  the  animal  dies,  the  blood 
is  non-coagulable.  Apparently  cells  of  nearly  all  types  can  be  de- 
stroyed b}^  trypsin,  which  may  cause  necrosis  in  one-fourth  hour ;  how- 
ever, spermatozoa  and  surface  epithelium  resist  strong  trypsin  solu- 
tions. Intravenous  injections  cause  death  with  lesions  in  the  heart 
muscle  and  severe  hemorrhages.  After  recover}^  from  one  injection 
of  trypsin  the  animal  is  temporarily  somewhat  more  resistant  to 
another  injection,  and  there  are  other  resemblances  to  anaphylactic 
intoxication  (Kirchheim -^).  Fiquet -*  also  observed  that  trypsin  and 
pepsin  rendered  the  blood  incoagulable,  but  after  some  time  the 
coagulability  of  the  blood  is  increased  and  thrombosis  is  frequent. 
Wells  -^  found  that  pancreatic  extracts  containing  veiy  active  trypsin 
and  lipase,  injected  intraperitoneally,  produced  an  acute  inflammatory 
reaction,  but  no  fat  necrosis.  Extracts  containing  active  lipase  and 
inactive  trypsin  were  less  toxic,  but  produced  fat  necrosis.  Extracts 
of  liver  and  blood  serum,  rich  in  lipase,  were  almost  without  effect  on 
dogs  and  cats.  Papain  was  found  to  be  much  more  toxic  than  any 
animal  enzyme,  causing  violent  local  hemorrhagic  inflammation. 
Schepilewsky  -^  also  found  papain  much  more  toxic  than  rennin  and 
pancreatin;  repeated  injection  of  the  two  latter  caused  amyloidosis 
in  rabbits.  Active  immunity  does  not  follow  repeated  injections  of 
papain.-^  Lombroso  -*  found  that  inactive  pancreatic  juice  was  much 
less  toxic  than  the  activated,  showing  that  it  is  the  trypsin  that  is  the 
important  toxic  agent.  He  also  found  that  succus  entericus  in  doses 
of  1  to  5  c.c.  is  toxic,  but  not  lethal  for  dogs.  Pancreatic  lipase  is 
hemolytic  (Noguchi-^)  if  activated  by  fats,  which  suggests  that  when 
this  enzyme  gets  into  the  blood  it  may  cause  hemolysis.  Hildebrandt  ^^ 
observed  that  enzymes  Avere  positively  chemotactic,  but  it  is  probable 
that  the  products  of  their  action  on  the  tissues  are  the  chief  chemo- 
tactic agents. 

The  enzymes  that  are  secreted  into  the  gastro-intestinal  tract  seem 
to  be  chiefly  destroyed,  but  part  is  eliminated  in  the  feces,  and  part 
that  is  absorbed  apparentl}^  reappears  in  the  urine  in  ver}^  small 
quantities.^^     Pepsin,  diastase,  and  rennin  all  have  been  found  in  nor- 

22  Ann.  d.  I'Inst.  Pasteur,  1001   (15),  737. 

23  Arch.  exp.  Patli.  u.  Pliarni.,  mil    (GO),  352;    1914   (78),  99;    1913    (74),  374. 

24  Arch.  d.  M^'d.  Kxpor.,  1899    (11),  145. 

25  Jour.  Med.  Hesearcli,  1903    (9),  92. 

26  Cent.  f.  Pakt..  1899   (25),  849. 
2TStenitzer,  Piocliom.  Zeit..  1908   (9),  382. 

2R  Al)8tract  in  Piuchoni.  Contralblatt,  1903   (1),  712. 

20  Pioclicni.  Zeit.,   1907    ((i),  185. 

so  Virchow's  Arch.,   1893    (131),  5. 

31  Falk  and  Kolicb,  Zeit.  klin.  'Mod.,  1909    (08),  15G. 


A\Ti-E\zy.][i:s  63 

mal  urine;  but  trypsin  is  present  chiefly  as  trypsinogen,  especially 
abundant  after  a  meat  diet.''-  Pepsin  and  rennin  enter  the  urine  as 
the  zymogens,  in  quantities  in  proportion  to  the  amount  in  the 
stomach,  and  are  absent  in  gastric  carcinoma  (Puld  and  Hirayama^^). 
During  resolution  of  pneumonia,  leucocytic  protease  may  appear  in 
the  urine  (Bittorf  ^*).  Ferments  injected  subcutaneously  seem  sel- 
dom to  be  eliminated  in  any  considerable  amounts  in  the  urine,  but 
Opie  ^^  has  demonstrated  the  presence  of  lipase  in  the  urine  in  pan- 
creatitis with  fat  necrosis.  Hildebrandt  was  able  to  prove  that  emulsin 
remained  active  for  at  least  six  houi's  after  it  was  injected  into  animals 
subcutaneous!}',  by  its  splitting  amygdalin  which  was  then  injected, 
the  CNH  liberated  by  the  cleavage  of  the  amygdalin  causing  death. 

ANTI-ENZYMES 

Injection  of  enzymes  into  animals  leads  to  the  appearance  of  sub- 
stances in  the  senim  of  the  animals  that  antagonize  the  action  of  the 
enzymes.^^  The  principles  involved  are  quite  the  same  as  in  the 
immunization  of  animals  against  bacterial  toxins  or  against  foreign 
proteins.  This  seems  to  have  been  first  observed  by  Hildebrandt, 
and  it  has  been  taken  up  extensively  in  recent  years  in  the  stud}'  of  the 
problems  of  immunity.  An  interesting  observation  that  was  made 
rather  early  in  these  studies  was  that  normal  blood-serum  possesses 
a  marked  resistance  against  the  action  of  proteolytic  enzymes,  not 
being  at  all  digested  by  dilutions  of  enzymes  that  will  rapidly  digest 
a  serum  that  has  been  heated.  This  property  seems  to  be  shared  by 
egg-white  ^^'^  and  by  the  tissues  and  organs  of  the  body  (Levene  and 
Stookey^^).  The  anti-enzyme  action  is  easily  destroyed,  by  heat  of 
about  70°,  by  the  action  of  dilute  acids,  and  even  by  prolonged  stand- 
ing. It  is  exerted  not  only  against  the  secreted  ^proteolytic  enzjones, 
pepsin  and  trypsin,  but  also  against  the  intracellular  enzymes  of 
various  organs. 

It  seems  highly  probable  that  the  resistajice  of  the  body  tissues  to 
digestion  by  their  own  enzymes  and  by  the  enzymes  of  one  another 
depends  in  some  way  upon  the  presence  of  anti-enzymes  in  the  cells 
;ind  tissue  fluids,  for  self -digestion  of  tissues  is  greatly  impeded  by 
serum.^^     Weiland^*'  has  demonstrated  that  certain  intestinal  worms 

32  V.  Schoenborn.  Zeit.  f.  Biol..  1010    (53),  386. 

33Berl.  klin.  Woch.,  1010   (47),  1062. 

3*  Dent.  Arch.  klin.  :\led..  1007    (91),  212. 

30  Johns  Hopkins  Hosp.  Bull.,  1902    (13),  117. 

37  According  to  Porter  (Quart.  .Joxir.  Exper.  Physiol.,  1910  (3),  37o)  enzymes 
in  contact  with  various  membranes  are  inactivated,  and  substances  appear  which 
are  strongly  inhibitivo  to  the  enzymes:  it  is  possible  that  this  effect  depends 
largely  on  zvmoids,  which  unite  with  the  substrate  and  deviate  the  enzymes. 

37aSugimoto.  Arch.  exp.  Path..  1013   (74),  14. 

38  Jour.  Medical  Pesearch.  1003   (10),  217. 

39  Wells,  Jour.  Med.  Pesearch,  lOOG    (10).  149. 

•to  Zeit.  f.  Biol..  1003  (44),  45:  see  also  Dastre  and  Stassano.  Compt.  Pend. 
Soc.  Biol.,  1003   (55),  130  and  254:  and  TTamill,  Jour,  of  Physiol.,  1906   (33),  470. 


64  ENZYMES 

contain  a  strong-  antitrypsin,  to  which  lie  attributes  their  ability  to 
live  bathed  in  pancreatic  juice  without  being  digested.*"'^  Similar 
properties  have  been  ascribed  by  other  observers  to  the  cells  of  the 
mucosa  of  the  stomach*^  and  intestine,  and  to  the  mucus  itself  (de 
Klug)/-  but  the  work  of  Bensley  and  Harvey  ^^  indicates  that  the  ab- 
sence of  free  acid  in  the  gland  cells  and  lumen  is  perhaps  the  chief 
protection  of  the  stomach  from  pepsin.  Kirchheim  **  holds  that  the 
intestines  are  protected  less  by  anti-enzymes  than  by  rapid  absorption 
and  removal  of  the  enzymes,  which  are  really  not  present  in  any  con- 
siderable excess  in  the  intestinal  contents.  The  anti-enzymes  seem 
only  to  inhibit  enzyme  action,  and  not  to  destroy  the  enzyme  itself.*"' 
Normal  anti-enzj-mes  do  not  seem  to  be  at  all  specific,  according  to 
V.  Eisler,*"  that  is,  human  serum  is  no  more  resistant  to  human  tryp- 
sin than  is  pig  serum — indeed,  it  is  less  so.*' 

Cathcart  **  found  that  antitrypsin  is  connected  ^\^th  the  ' '  albumin 
fraction"  of  the  serum,  i.  e.,  the  fraction  precipitated  between  half 
and  full  saturation  with  ammonium  sulphate.  Globulins  do  not  pos- 
sess this  action,  but  they  are  not  easily  digested.  Antitrypsin  is 
found  in  all  varieties  of  serum,  and  is  little  or  not  at  all  specific.  It  is 
destroyed  by  65-70°  C.*"  for  one-half  hour,  but  retains  its  anti-enzy- 
matic activity  after  drying,  and  is  equally  effective  against  all  sorts 
of  proteins.  The  normal  anti-tryptic  activity  decreases  during  fast- 
ing and  increases  during  digestion  (Rosenthal  ^")  ;  it  is  increased 
during  pregnancy  ^"^  and  the  blood  of  the  fetus  shows  less  than  that 
of  the  mother.  Normal  antitrj^psin  unites  with  trypsin  according  to 
the  law  of  multiple  proportions  (^leyer)  and  the  reaction  is  not  re- 
versible (Rondoni).  It  is  found  in  the  urine,  and  in  inflammator}' 
exudates,  but  not  in  normal  serous  fluids,  and  it  resists  putrefaction. 
Normal  serum  does  not  seem  to  inhibit  the  enzymes  which  act  upon 
purines.     Fuld  and   Spiro  '^^   found   that   the   natural   antirennin   of 

4oa  Burge  (Jour.  Parasitol.,  1915  (1),  179)  suggests  that  the  protection  of 
parasites,  and  perhaps  of  the  alimentary  epithelium,  depends  on  tlie  active 
oxidative  pro])erties  of  tliese  tissues  destroyinsr  tlie  en/.vmes. 

41  See  Blum  and  Fuld,  Zeit.  Iclin.  Med., 'lOOG  (fiS),  505;  LangensldoUl.  Skand. 
Arch.  Physiol.,  1914   (31),  1. 

•»2  Arch,  internat.  d.  phvsiol.,  1007    (5),  297. 

■»3  Biological  Bulletin,  1912    (2.3),  225. 

44Arch.  exp.  Bath.  u.  Pharm.,  1912    (71),  1 

45  l^ayliss  and  Starling  (Jour,  of  Physiol..  1905  (.32).  129;  and  Meyer.  Biochem. 
Zeit.,  1909  (23),  OS.  o])pose  the  view  of  Dele/enne  that  the  antitryptic  action  of 
the  hlood  is  due  to  an  antikinase.  and  believe  the  antibody  acts  upon  trypsin. 

■»«  Ber.  d.  Wien.  Akad.,   1905    (104),  119. 

4"  This  is  contradicted  by  Claessncr;  llofmeister's  Beitriige,  1903   (4),  79. 

48  .Tour,  of  Phvsiol.,  1904  (31),  497;  also  see  Kiimmercr  and  Aubrv .  l?i(iclicni. 
Zeit.,  1913    (48),'  247. 

40  Unless  otherwise  specified,  all  tem])craturcs  are  given  according  to  the  Centi- 
grade scale. 

->o  Folia  Serologica.  1910  (0),  2S5 ;  also  Fran/,  and  Jarisdi,  Wien.  klin.  Woch.. 
1912    f25).   1441. 

noaSee  Franz,  Arch.  f.  CJvn.,  1914    (102).  579. 

51  Zeit.  f.  phvsiol.  Chem."  1900   (31).  132. 


A\Tf-K\Z)]n:,<<  65 

normal  horse  serum  is  in  the  pseu(l()<il()l)ulin  fraction.  Since  acids 
destroy  the  anti-enzyme  property  of  the  serum,  it  is  not  effective 
against  pepsin-IICl  mixtures.  Against  trypsin,  however,  it  is  very 
effective.  Zniiz  ■'-  states  that  normal  serum  acts  more  upon  entero- 
kinase  than  upon  try!)sin,  and  believes  tlmt  the  inliibition  depends 
upon  colloids  which  modify  surface  tension  and  adhere  to  the  pro- 
teins. Red  corpuscles  and  living  unicellular  organisms,  including 
bacteria,  are  likewise  resistant  to  trypsin,  and  normal  serum  also 
seems  to  contain  an  antirennin."'?  Fresh  and  inactivated  serum  will 
prevent  pepsin  from  digesting  protein,  but  this  is  not  due  to  a  true 
antipepsin,  according  to  Hamburger.^*  Oppenheimer  and  Aron  ^^' 
consider  it  probable  that  the  resistance  of  normal  serum  to  trypsin  di- 
gestion depends  upon  the  configuration  of  the  protein  molecules,  which 
perhaps,  when  in  fresh,  uninjured  condition,  present  no  suitable  sur- 
faces for  attack  by  the  ferment. 

Jobling  '^■'"-  and  his  co-workers  have  advanced  evidence  that  the  nor- 
mal antiprotease  action  of  serum  depends  on  the  lipoids  of  the  serum, 
which  vary  in  activity  directly  with  the  degree  of  uusaturation ;  there- 
fore they  were  able  to  decrease  the  antiferment  action  of  serum  by 
extracting  the  lipoids  with  fat  solvents  (and  to  restore  the  activity 
by  replacing  the  lipoids),  or  by  saturating  the  double  bonds  of  the 
fatty  acids  with  halogens,  or  by  modifying  the  degree  of  dispersion 
of  the  lipoids.  Soaps  of  saturated  fatty  acids  do  not  inhibit  serum 
protease. 

Opie  ■'''  has  found  that  the  serum  of  inflammatory  exudates  con- 
tains an  anti-enzymatic  substance,  destroyed  at  75°,  and  by  acids; 
it  is  not  present  in  normal  cerebrospinal  fluid,  but  appears  here  as 
in  other  serous  cavities  during  inflammation  (Dochez).'^"  Antitrypsin 
has  also  been  found  in  pathological  urines  (v.  Sclioenborn).^^ 

The  power  of  the  blood  semm  to  inhibit  the  activity  of  trypsin  and 
leucocytie  protease  has  been  found  to  vary  greatly  in  disease,  and,  as 
having  diagnostic  possibilities,  this  property  has  been  considerably 
investigated.^^  It  is  especially  increased  in  conditions  associated  with 
cell  destruction,  such  as  pneumonia  and  cancer,  which  suggests  that 

52  JTem.  Acad.  roy.  med.  Bclgiqiie,  1909   (20).  fase.  5. 

53Czapek  (Ber.  Deut.  botan.  Gesell.,  1903  (21),  229)  states  lliat  anti-oxidases 
occur  normally  in  certain  plants,  strongly  specific  aprainst  tlie  oxidase  of  the 
same  plant  species. 

54  .Jour.  Exper.  'Sled.,  1911  (14),  .5.35;  Arcli.  Tnt.  ^led.,  1915  (16),  .350.  There 
seems  to  be  no  relation  between  the  antipeptic  and  antitrvptic  powers  of  sera 
(Rubinstein,  Ann.  Inst.  Pasteur.,  1913    f27).  1074). 

5- Hofmeister's  Beitriige,  1903    (4),  279. 

55a  Series  of  articles  in  Jour.  Exper.  ^led. :  also  review  in  .Tour.  T.al).  and  Clin. 
Med..  1915    (1),  172.     See  also  Zeit.  Tmmunitiit.,  1914   (23),  71. 

50  Jour.  Exp.  Med..  1905   (7).  31fi. 

57, Jour.  Exper.  Med.,  1909   (11),  718. 

5sZeit.  f.  Biol.,  1910   (53),  386. 

59  For  literature  and  review  see  Wiens,  Erjiebnisse  Phvsiol..  1911  (15).  1; 
Weil,  Arch.  Int.  Med.,  1910   (5),  109;  Meyer,  Folia  Serologica,  1911    (7),  471. 

5 


66  ENZYMES 

the  increased  aiititryptic  activity  results  from  the  formation  of 
specific  antibodies  for  the  intracellular  proteases  liberated  during  the 
disease,  but  as  yet  this  has  not  been  satisfactorily  established,  so  we 
do  not  know  whether  the  "antitrypsin  reaction"  depends  upon  an 
antibody  for  trypsin  or  upon  some  entirely  different  factor.  In 
cachexia  the  inhibiting  effect  of  the  serum  is  especialh'  marked  and 
it  is  therefore  usually  pronounced  in  cancer,  but  the  increased  inhibi- 
tion is  sometimes  absent  in  cancer  (10  per  cent,  of  all  cases)  and  often 
present  in  other  conditions,  so  that  the  positive  diagnostic  value  is 
slight.  It  may  also  be  present  without  cachexia  and  often  seems  to 
parallel  the  number  of  leucocytes  in  the  circulating  blood.  Sarcoma 
shows  it  less  than  carcinoma,  while  in  exophthalmic  goitre  and  tuber- 
culosis an  antitryptic  increase  is  said  to  be  quite  constant  (Waelli).*"^ 
As  normal  serum  contains  a  tryptic  enzyme  as  well  as  a  substance 
inhibiting  trypsin,  the  antitryptic  activity  is  at  most  but  a  measure  of 
the  difference  between  these  (Weil),  and  might  depend  on  either 
lowered  trypsin  or  increased  antitrypsin  content.  Doblin  *'^  and  many 
others  believe  with  Jobling  that  the  active  agent  is  not  a  true  immune 
antibody,  but  as  yet  general  agreement  has  not  been  reached  on  this 
point  (see  Meyer).  Kirchheim  ^^'^  has  found  that  the  union  of  trypsin 
and  antitrypsin  does  not  follow  the  physico-chemical  laws  of  a  true 
antigen-antibody  reaction.  Rosenthal  has  advanced  evidence  to  sup- 
port the  hypothesis  that  the  presence  of  products  of  protein  cleavage 
in  the  serum  is  responsible  for  the  antitryptic  action,  but  this  has  not 
been  confirmed.  Attempts  have  been  made  to  regulate  suppurative 
processes  by  the  introduction  of  either  leucocytic  proteases,  or  anti- 
protease  in  the  form  of  active  serum  (see  Wiens  •''''').  Whether  anti- 
protease  can  be  specifically  developed  by  immunizing  with  leuco- 
protease  is  a  matter  of  disagreement,**-  but  no  increase  of  antiprotease 
follows  the  enormous  destruction  of  leucocytes  caused  by  injecting 
tliorium-A'."-' 

The  anti-enzymatic  property  obtained  in  the  serum  by  injecting 
enzymes  into  animals  differs  from  that  normally  present  in  the  senim 
in  many  ways.  It  may  be  made  much  stronger  than  it  ever  is  in 
normal  serum,  and  against  many  varieties  of  enzymes  for  which  an 
anti-enz.yme  docs  not  naturally  exist.  Especially  important  is  the 
fact  that  it  is  highly  specific  (v.  Eisler)  ;  serum  of  an  animal  immu- 
nized against  dog  trypsin  will  show  a  much  greater  effect  against  dog 
trypsin  than  it  does  against  tryi)sin  from  other  animals.  This  fact 
permits  us  to  distinguisli  between  enzymes  of  ap]iarently  similar  na- 
ture but  of  different  origin,  and  ])roves  that  they  have  a  structure  at 

60  Mitt.  Crenz.  Mod.  u.  Cliir..  1912    (25),  1S4. 
11  Zeit.  f.  Iminunitiit,  1009   (4),  229. 
Ola  Arch.  oxp.  Patli..  191.3   (7.3),  1.39. 
«2  See  Bradley,  Jour,  llyg.,  1910   (12),  209. 
03  G.  Rosenow  and  Fiirb'er,  Zcit.  cxp.  Mod.,  1914   (3),  377. 


RESEMBLANCES  OF  ENZYMES  AND  TOXINS  G7 

least  ill  somo  respects  different  from  one  another,  since  they  are  com- 
bined by  different  antibodies.  Apparently  that  element  of  the  en- 
zymes which  detei-mines  their  action  on  specific  substances  is  involved 
in  their  antigenic  properties,  since  antiproteases  will  not  inhibit  dias- 
tase or  lipase.  This  specificity  is  limited,  however,  for  the  anti-en- 
zymes for  leucocytic  and  pancreatic  proteases  are  said  to  be  iden- 
tical.^* Artificial  immune  serum  is  said  to  have  been  obtained  aj^ainst 
trypsin,  pepsin.^*  lipase,  emulsin,''^  autolytic  enzymes,  laccase,  amyl- 
ase, invertin,  diastase,  tyrosinase,  urease, '^"'^  rennin,  catalase,  and 
fibrin  ferment.*'*  By  immunization  against  bacteria  an  immunity 
against  their  proteolytic  enzymes  is  also  obtained,"^  which  is  inde- 
pendent of  and  different  from  antitrypsin,  being  especially  in  the 
globulin  fraction,  while  the  antibody  for  pancreatic  trypsin  is  chiefly 
in  the  albumin  (Kammerer ''^).  From  the  work  of  Kirchheim  and 
Reinicke  *^-^  it  seems  probable  that  the  increased  resistance  following 
immunization  with  trypsin  is  simply  an  increase  in  nonspecific  resist- 
ance, such  as  follows  injection  of  peptone  and  many  other  poisonous 
substances.  There  is,  indeed,  a  growing  suspicion  that  much  of  the 
evidence  of  specific  antibody  formation  for  enzymes  must  be  revised. 
Resemblances  of  Enzymes  and  Toxins. — As  can  be  seen  from  the 
above  statements,  the  enzymes  behave  in  many  respects  like  the  tox- 
ins, both  in  their  manner  of  acting  upon  other  substances  and  in  the 
reaction  they  produce  when  introduced  into  the  bodies  of  animals. 
As  Oppenheimer  says,  "the  bonds  between  enzymes  and  toxins  are 
drawing  closer  and  closer."  According  to  some  experiments,  the 
enzymes  behave  much  as  if  they  possessed  a  haptophore  and  a  toxo- 
phore  group,  the  former  of  which  combines  with  the  substance  that  is 
to  be  acted  upon;  and  immunity  appears  to  be  produced  by  the  de- 
velopment of  receptors  that  combine  the  haptophore  groups,  these 
receptors  constituting  the  antiferments.  There  is  abundant  evidence 
of  a  toxin-like  structure  in  enzymes,  from  the  numerous  observations 
on  the  formation  of  "zymoids"  which  can  neutralize  anti-enzymes  or 
combine  with  the  substrate,  although  no  longer  active  as  enzymes. 
The  oxidizing  enzymes  especially,  with  their  complex  relationship  of 
substrate,  combining  body  (peroxides)   and  enzyme,  present  striking 

64  .Jochmann  and  Kantorowiez,  IMiinch.  med.  Wocli.,  1908.  (5.5),  728. 

65  Bayliss  (.Tour,  of  Physiol.,  1912  (4-3),  455)  was  unable  to  obtain  anti- 
emulsin,  and  Pozerski  (Ann.  Inst.  Pasteur,  1909  (2.3),  20."))  failed  to  ol)tain  anti- 
papain,  but  positive  results  are  reported  bv  v.  Stenitzer  (Biochem.  Zeit..  1908  (9), 
382). 

63a  .Tacoby  says  that  the  disappearance  of  mease  from  the  blood  after  repeated 
injection  does  not  depend  on  the  formation  of  an  anticnzvme  (Biochem.  Zeit., 
1916   (74),  97). 

66  For  a  review  of  mucli  of  the  earlier  literature  on  this  subject  see  Schiitze, 
Deut,  med.  Woch.,  1904   (30),  308. 

6"  V.  Dunpern,  ^Fiinch.  med.  Wochenschr.,  1898  (45),  1040;  Bertiau,  Cent.  f. 
Bact.,  1914   (74),  374. 

68  Deut.  Arch.  klin.  Med..  1911    (103).  341. 
68a  Arch.  exp.  Path.,  1914   (77),  412. 


68  EXZYMEfy 

analogies  to  immune  reactions  (^loore"^"),  and  the  proteolytic  sub- 
stances of  the  blood  resemble  the  lysins  in  certain  respects  (Dick).'° 
Enzj'uies  and  toxins  also  resemble  one  another  in  being  readily  ab- 
sorbed by  membranes,  precipitates,  and  highly  developed  surfaces  in 
general.'^  Finally,  there  is  much  reason  to  believe  that  the  hemo- 
lytic toxin  of  cobra  venom  is  a  lipase,  which  acts  by  splitting  lecithin 
into  hemolytic  substances  (Coca)."^* 


THE  INTRACELLULAR  ENZYMES  "- 

Until  a  recent  time  our  knowledge  of  enzymes  in  the  animal  body 
was  limited  to  those  present  in  the  digestive  secretions.  AVith  few 
exceptions  these  are  without  influence  in  pathological  processes,  since 
they  seem  to  be  but  little  absorbed,  and  rarely  enter  the  blood  or 
tissues  in  any  other  way.  But  with  the  more  recently  disclosed  intra- 
cellular enzymes,  many  of  which  are  present  in  every  cell,'^  the  rela- 
tion to  pathology  is  very  intimate.  These  intracellular  enzymes,  as 
we  now  know  them,  and  their  chief  properties,  are  as  follows : 

OXIDIZING  ENZYMES  '* 

Although  oxidation  of  organic  compounds  is  the  chief  source  of 
energy  in  the  animal  body,  yet  the  way  in  which  it  is  accomplished 
is  very  little  understood.  We  only  know  that  it  is  brought  about 
within  the  cells,  and  that  substances  that  outside  the  body  are  oxidized 
with  difficulty,  are  completely  oxidized  to  carbon  dioxide  and  water 
within  the  cells,  and  that  this  is  done  with  just  such  a  degree  of  rapid- 
ity that  the  heat  produced  is  in  exactly  the  amount  necessary  for 
the  wants  of  the  body.  There  can  be  little  question  that  this  oxida- 
tion is  accomplished  through  catalytic  agents  acting  within  the  cells, 
and  certain  of  them  have  been  placed  in  a  condition  permitting  of 
study.  As  yet  their  exact  relations  to  intracellular  oxidation  are 
not  clearly  defined,  but  for  the  present  they  may  be  grouped  pro- 
visionally as  oxidizing  enzymes.     That  some  of  them  are  highly  specific 

«n  TJiofhom.  Jour.,  1909    (4),  1G5. 

70  .Tour.  Infectious  Diseases,  1911   (9),  282. 

71  See  Porter,  Quart.  Jour.  Exp.  Phvsiol.,  1010   (3),  ,375. 
7ia,Jour.  Infect.  Dis..  1915   (17).  .351. 

72  See  Vernon,  Ergebnisse  d.  Physiol.,  1910  (9),  13S;  also  liis  monograph, 
"Intracellular  Knzvmes."'  London,  1908. 

73irorlitzka  (Arch.  kal.  hiol.,  1907  (48).  119)  and  otlicrs  have  shown  that 
the  difTcrent  enz>ines  ajipear  one  hy  one  in  the  d(>v('lo])nHMit  of  the  ovum.  Tlieir 
activity  is  modified  considerahly  by  infections  (Sieher.  Piocliem.  Zeit..  1911  (32). 
108)   and  other  diseases   (Grossinaiin.  (7)(V7..  1912   (41).  181). 

74  f"om})lete  1)il)iio<rraphv  and  exliaustive  discussion  bv  Kastle.  liuli.  Hvpienic 
Lab.,  No.  59;  by  Loele,  Eifrob.  allfj.  Path.,  1912  (10,  Pt.'2).  700:  and  by  Rattelli 
and  Stern.  Erpebnisse  d.  Pliysiol.,  1912  (12).  90.  Concerninsr  the  cliemistry  of 
vital  oxidations  see  Dal<in.  "Oxidations  and  Reductions  in  the  Animal  Body," 
!Mono{rraplis  on  Piocliemistry.  London.  1912.  Good  review  l)y  v.  I'iirtli.  "Chem- 
istrv  of  Metabolism."  Cliaps.  22  and  23;  translated  bv  A.  J.  Smith.  Pliiladclphia. 
1910. 


CATALA8E  60 

is  shown  by  those  disorders,  such  as  alkaptonuria  and  diabetes,  in  which 
tlio  body  loses  the  power  to  oxidize  a  certain  cliemical  substance  while 
retaining'  the  normal  power  to  oxidize  innumerable  other  substances. 
Aecordinof  to  Lillie  ' '"  the  oxidative  processes  in  cells  take  place  most 
actively  in  relation  to  the  membrane  surfaces  (or  phase  boundaries)  of 
the  cells.  Of  the  oxidizing  enzymes  as  yet  identified  none  can  be  con- 
sidered as  of  importance  in  the  energy-producing  oxidations  of  the 
body  (iiattelli  and  Stern),  all  the  enzymes  of  this  class  yet  known  be- 
ing apparently  concerned  with  less  essential  oxidizing  processes;  it  is 
indeed  possible  that  the  essential  oxidation  of  food-stutt's  may  not  be 
dependent  on  enzymes  (Engler  and  Herzog).'^-''  An  agent  accelerating 
the  essential  oxidizing  activities  of  the  tissues  has  been  described  by 
Battelli  and  Stern  '"^  under  the  name  of  pneUi,  and  an  antipneuniin 
which  holds  it  in  check.  Closely  related  to  the  oxidizing  enzymes  is — 
Catalase. — It  has  long  been  known  that  most  enzymes  possess  the 
power  of  decomposing  hydrogen  peroxide,  with  liberation  of  oxygen ; 
but  it  was  not  until  1901  that  it  was  finally  demonstrated  by  Loew 
that  this  property  was  due  to  a  separate  enzyme  and  was  independent 
of  the  specific  properties  of  the  various  other  enzymes.  This  ferment 
is  very  wide-spread,  and  so  is  generally  obtained  along  with  the  other 
enzymes  when  attempts  are  made  to  isolate  them  from  the  cell.  It 
was  named  catalase  by  Loew,  and  he  described  two  fonus,  a-catalase, 
which  seems  to  be  a  nucleoprotein,"'*''  and  (i-catalase,  which  has  the 
properties  of  an  albumose.  It  has  been  demonstrated  by  Bach  and 
Chodat  that  peroxides  are  contained  in  plant  cells,  and  they  also  occur 
in  animal  cells.  According  to  Golodetz  and  Unna  "^  the  catalases  are 
held  in  the  cytoplasm  of  the  cells  while  the  peroxidases  are  in  the 
nucleus.  Just  what  function  the  catalase  performs  is  at  present  merely 
a  matter  of  speculation,  but  that  it  serves  an  important  purpose  is 
indicated  by  the  observation  of  Burge  ^""^  that  the  amoimt  of  catalase 
in  muscles  varies  directly  with  their  activity.  Loew  considers  that  it 
destroys  peroxides  formed  in  metabolism,  which  are  very  poisonous  to 
cell  life.  Shaffer  has  found  evidence  that  under  the  influence  of  cata- 
lase the  oxygen  liberated  is  in  the  molecular  form,  O^,  and  therefore 
relatively  inert;  whereas  when  peroxides  spontaneously  decompose, 
they  liberate  atomic  oxygen  which  is  an  active  oxidizing  agent.  He 
found  that  uric  acid  is  oxidized  by  peroxide  of  hydrogen,  but  when 
catalase  is  present,  this  oxidation  is  prevented.  According  to  this 
the  function  of  catalase  is  rather  to  prevent  dangerous  forms  of  oxida- 
tion than  to  help  in  normal  oxidative  processes.  P^'or  the  present, 
however,  nothing  can  be  said  positively  on  this  subject. 

T4a  Jour.  Biol.  Chem.,  1913   (15),  2.37. 

75Zeit.  physiol.  Chem.,  1009   (59),  327. 

76Biocliem.  Zeit.,  1911    (33),  315;   1911    (36),  114. 

76a  Xot  corroborated  tjv  Waentifj  and  Gierisch,  Fermentforsch.,   1914    (1).   165. 

77Berl.  klin.  Woch.,  1912   (49),  11,34. 

"aAmer.  Jour.  Pliysiol.,  1910    (41),  153;   1917    (42),  373. 


70  EXZYME& 

Occurrence  of  Catalase  wider  Normal  and  Pathological  Condi- 
tionsJ~^ — Battelli  and  Stern  found  that  the  catalytic  power  of  the  tis- 
sues endures  man}'  hours  after  dyath.  Its  abundance  is  ditferent  for 
different  organs  of  the  same  animal,  but  remarkably  constant  for  the 
same  organ  in  the  same  species.  In  general  the  order  in  decreasing 
strength  is:  liver,  kidney,  blood,  spleen,  gastro-intestinal  mucosa,  sal- 
ivary glands,  lung,  pancreas,  testicle,  heart,  muscle,  brain ;  but  this 
order  varies  in  different  species.  Catalase  is  abundant  even  in  the 
early  embrj'o  (Mendel  and  Leavenworth)  and  in  sea  urchin  eggs  it 
increases  rapidly  after  they  are  fertilized  (Lyon).'*  Leucocytes  con- 
tain little,  most  of  that  in  the  blood  being  in  the  stroma  of  the  red 
blood-corpuscles.  The  body  fluids  contain  little  or  none.  Injected 
intravenously,  catalase  (of  the  liver)  is  destroyed  rapidly,  and  does 
not  appear  in  the  urine ;  it  does  not  cause  any  toxic  effects,  nor  does 
it  increase  resistance  to  poisoning  by  venoms.  The  tissues  also  con- 
tain anti-catalases,  and  still  further  a  substance  which  protects  the 
catalase  from  the  anti-catalase ;  this  protective  substance  is  called  the 
philocatalase  by  Battelli  and  Stern. 

The  gas  evolved  by  the  action  of  pus  on  H0O2  was  found  by  ]Mar- 
shall ''°  to  be  pure  oxygen,  each  c.c.  of  a  certain  sample  of  pus  exam- 
ined liberating  133.9  c.c.  of  gas.  The  active  constituent  of  pus,  he 
states,  is  contained  in  the  serum  and  not  in  the  corpuscles.  Substances 
decomposing  HoOj  have  been  found  also  in  bacterial  cultures,  first  by 
Gottstein,  and  later  in  the  cell  juices  expressed  from  tubercle  bacilli 
by  Hahn.  Loewenstein  ^°  found  an  enzyme  agreeing  with  catalase  in 
filtered  bouillon  cultures  of  diphtheria  bacilli  and  staphj-lococci,  but 
not  from  tetanus,  typhoid,  and  colon  bacilli  or  cholera  vibrios;  the 
catalase  is  quite  distinct  from  the  toxin.  He  also  found  that  the  ad- 
dition of  H2O2  to  a  diphtheria  toxin-antitoxin  mixture  destroyed  the 
toxin,  leaving  the  antitoxin  free.  A  similar  destruction  of  tetanus 
toxin  by  peroxides,  first  demonstrated  by  Sieber,  can  occur  without 
the  catalase. 

Winternitz  ^'  and  his  associates  have  made  extensive  studios  of  the 
catalase  activity  of  the  blood  and  tissues  in  disease.  They  found  that 
all  tissues  have  reduced  catalase  activity  in  chronic  nephritis,  in  pro- 
portion to  the  severity  of  the  condition,  and  experimental  nephritis 
in  animals  has  the  same  effect;  the  blood  shows  gi'oat  reduction  in 
catalase  in  uremia,  and  a  less  reduction  witli  less  severe  nephritic 
manifestations:  Eclampsia  shows  little  or  much  reduction  of  catalase 
in  the  blood  in  proportion  to  the  amount  of  renal  involvement ;  normal 
pregnancy  and  labor  have  no  effect.     Anemia  is  associated  with  irregu- 

7Vb  Conooniinfr  the  oatalaso  of  lower  animals   see  Zicgor.   Rioclicm.   Zcit.,   1915 
(69),  .39. 
78Amer.  .lour.  I'livsiol.,  l!)Od    (Sf)).  inO. 
70  Univ.  of  Pcnn.  Med.  Bull.,  1902   (15),  366. 
RoWien.  klin.  Woeh.,  100.1   (10),  1.393. 
81  Review  in  Aroli.  Int.  Med.,  1911    (7),  624. 


oxiDi'/.iSd  i:\/.)  ][j:s  71 

lar  decrease  in  eatalase,  including  primary  anemias  and  the  secondary- 
anemias  of  typhoid  and  pneumonia;  cardiac  disease  has  no  effect  if 
tlie  kidneys  are  normal.  Acute  peritonitis  causes  a  rise  in  l)lood 
eatalase;  diabetes,  leukemia  and  jaundice  were  without  effect.  In 
hyperthyreosis  the  eatalase  tends  to  increase,  in  hypothyreosis  to  de- 
crease; complete  removal  of  the  thyroid  causes  a  decrease  which  disap- 
pears on  feeding  thyroid.  Intravenous  injection  of  salts,  acids  and 
alkalies  decreases  the  catalytic  activity  of  the  blood.  Normal  indi- 
viduals show  considerable  variations  in  the  eatalase  activity  of  the 
blood,  but  for  each  individual  it  is  remarkably  constant ;  age  has  very 
little  influence.  In  the  tissues  post  mortem  change  causes  but  slight 
reduction  in  eatalase.  Extirpation  of  large  amounts  of  kidnc}'  or  liver 
tissue  has  little  effect,  but  removal  of  the  spleen,  ovaries  or  testicles 
causes  a  transient  decrease  in  the  eatalase  of  the  blood.  If  the  red 
corpuscles  are  prevented  from  laking,  the  eatalase  activity  manifested 
by  the  blood  in  vitro  is  reduced  (Strauss)  ^~  and  iodides  increase  the 
eatalase  activity  of  the  blood.  Catalase  and  anticatalase  have  been 
found  in  pathological  urine,  in  both  acute  and  chronic  nephritis 
(Primavera).*^  Kahn  and  Brim  ^^'^  also  found  traces  of  catalase  in 
normal  urine,  greatly  increased  in  urine  containing  blood,  bile  or  ace- 
tone, normal  in  cancer,  high  in  diabetic  acidosis,  Hodgkin's  disease, 
septic  infections  and  typhoid.  Grossman  ^*  found  that  bacterial  poi- 
sons generally  increase  the  catalase  content  of  the  various  viscera,  and 
Rosenthal  *^  observed  a  great  decrease  in  the  liver  and  blood  of  mice 
receiving  intraperitoneal  inoculations  of  cancer.  The  catalase  activity 
of  the  non-cancerous  organs  of  cancer  patients  is  not  affected,  except 
slightly  lowered  by  cachexia  (Col well)  ^®;  however,  the  liver  tissue 
between  cancer  nodules  may  show  less  catalase  than  normal  liver. ^*''* 
True  Oxidizing  Enzymes. — AVhile  it  is  by  no  means  certain  that 
catalase  is  active  in  causing  intracellular  oxidations,  there  are  other 
enzymes  or  enzyme-like  substances  that  come  more  properly  under  the 
head  of  oxidases  or  oxidizing  enzymes.  Battelli  and  Stern  contend 
that  the  only  real  oxidases  which  have  yet  been  completely  established 
are:  1.  Polyphenoloxidases  (oxidizing  phenols  and  their  amino  com- 
pounds, but  not  tyrosine)  ;  2.  Tyrosinase;  3.  Alcohol  oxidase;  4.  Xan- 
thine oxidase ;  5.  Uricase.  Chodat  and  Bach  believe  that  the  enzymes 
which  are  designated  above  as  polyphenoloxidases  have  a  complex 
structure,  consisting  of  peroxidase  and  oxygenase.  Mathews  **"^  holds 
that  "under  the  term  oxidases  there  have  been  confused  two  classes 

82  Bull.  .Johns  Hopkins  Hosp.,  1912   (23),  120. 

83  Riforma  Med..  1906    (12),  1206. 
83aAmer.  .Jour.   Obst..   1915    (71).  39. 
84Biochem,   Zeit..    1912    (41),    ISl. 
85Deut.  med.  Woch..  1912    (38),  2270. 

86  Arch.  Middlesex  Hosp.,  1910   (19),  64. 

86a  Blumenthal   and   Brahn,   Zeit.  KrebsforBcli.,   1910    '8),   436. 

86b  Jour.  Biol.  Chem.,  1909    (6),  1. 


72  ENZYMES 

of  substances,  one  which  activates  the  oxygen;  the  other  the  more 
important  class,  which  activates,  by  dissociation,  the  reducing  sub- 
stances. The  latter  are  specific,  the  former  not."  This  view  has 
received  support  by  Bach. 

Peroxidase. — This  name  is  given  to  an  enzyme  that  is  believed  to 
cause  oxidation  by  activating  peroxides,  and  is  quite  distinct  from 
catalase  and  from  the  other  oxidases.  The  peroxide  on  which  it  chiefly 
acts  in  the  cell  is  supposed  to  be  the  so-called  "oxygenase." 

Oxygenase. — This  can  also  act  as  an  oxidizer  independent  of  the 
peroxidase,  in  the  presence  of  certain  manganese  compounds.  Loeven- 
hart  and  Kastle  question  the  true  enzyme  nature  of  this  and  other 
' '  oxidases, ' '  which  they  look  upon  as  organic  peroxides,  behaving  like 
other  peroxides  rather  than  as  catalyzers.  Practically  the  existence 
of  these  bodies  is  demonstrated  by  their  power  to  turn  tincture  of 
guaiac  blue,  and  they  are,  therefore,  present  in  pus. 

Von  Fiirth  '"*  sums  up  the  situation  in  these  words :  "In  the  tis- 
sues active  catalytic  agents,  the  peroxidases,  are  widely  distributed; 
which  seem,  just  like  the  coloring  matter  of  the  blood,  to  be  capable 
of  conveying  the  oxygen  from  peroxides  to  very  readily  oxidizable 
substances.  We  find  too  in  the  statements  bearing  upon  the  oxy- 
genases, the  aldehydases  and  indophenoloxidases,  occasion  for  .assum- 
ing that  there  are  substances  in  the  tissues  charged  with  oxygen  w^hich 
are  able  to  give  this  off  to  easily  oxidizable  matter ;  and  these  we  may 
in  a  measure  regard  as  peroxides.  But  that  is  all.  We  do  not  know 
whether  the  peroxidases  are  ferments  or  not." 

By  their  conception  of  oxygenase  and  peroxidase  Chodat  and  Bach 
would  displace  entirely  the  idea  of  enzymes  oxidizing  directly,  the 
true  "oxidases,"  which  they  consider  mixtures  of  oxygenase  and 
peroxidase.  There  have  been,  in  any  event,  a  number  of  ferments 
described  that  seem  to  possess  distinct  oxidative  powers.  As  each 
is  quite  specific  in  its  action,  oxidizing  but  one  substance,  or  one 
group  of  related  substances,  they  are  generally  designated  by  the 
name  of  the  substances  upon  which  they  act.  INIost  studied  of  these 
are  aldehydase  and  tyrosinase. 

Aldehydase,**"  which  is  characterized  by  oxidizing  aldehydes,  par- 
ticularly salicyl-aldehyde.  According  to  Jacquet,  this  enzyme  is  so 
intimately  bound  with  the  cell  that  it  cainiot  be  obtained  in  extracts 
until  after  the  cells  are  dead,  but  is  present  in  expressed  cell-,iuices. 
It  can  be  isolated  by  the  usual  methods,  is  destroyed  by  boiling,  acts 
best  when  no  free  oxygen  is  present,  and  its  action  is  inhibited  by 
CNII.  It  has  been  demonstrated  in  nearly  all  organs  and  tissues 
exce])t  pancreas,  muscle,  marrow,  and  mammary  gland ;  it  is  present 
in  the  blood  in  snudl  aiiKiunts.  but   not  at   all  in  the  l)ile.     It  is  most 

*>"  BattclH  and  Stern  do  not  include  aldeliydai^o  anion<,'  tin-  oxidizinj;  cn/ynips. 
on   the  ground  tliat  its  action  is  not  oxidative  but  hydrolytic. 


oxii)i7A\<;  j:\zy ]n:s  73 

abundant  in  the  liver''''  and  si)l(M'n,  and  is  )»rcsriii  in  |)i<f  (■inl)ryos, 
9  cm.  lony,  but  not  in  those  2-:}  em.  Ionj>'.  .Jacoby  luis  ol)tained  a  body 
with  the  properties  of  aldehydase  which  diil  not  <^ive  protein  re- 
actions. It  is  a  true  enzyme,  since  it  oxidizes  aldeliydes  without  itself 
being  used  up.  Its  range  of  action  is  limited,  for  Jacoby  found  it 
without  effect  upon  acetic  acid  and  stearic  acid. 

Tyrosinase. — This  enzyme,  which  is  found  both  in  animal  and  plant 
tissues,  is  particularly  interesting  in  relation  to  the  formation  of 
pigments.  Bertrand  found  that  the  transformation  of  the  juice  of 
lac-yielding  plants  into  the  black  lacquer  was  brought  about  by  the 
action  of  an  oxidizing  ferment,  laccase,  upon  an  easily  oxidized  sub- 
stance, laccol,  which  is  a  member  of  the  aromatic  series.  He  later 
found,  in  a  number  of  plants  an  enzyme  acting  on  tyrosine,  distinct 
from  the  laccase,  which  he  named  tyrosinase.  Biederman  later  found 
tyrosinase  in  the  intestinal  fluid  of  meal  worms,  v.  Fiirth  and 
Schneider  found  a  similar  enzyme  in  the  hemolymph  of  insects  and 
arthropods,  which  explains  its  darkening  when  exposed  to  air.  This 
enzyme,  as  obtained  from  different  sources,  is  not  always  specific  for 
tyrosine,  frequently  oxidizing  other  substances.  As  yet  the  chemical 
processes  and  end  results  of  the  oxidation  of  tyrosine  by  tyrosinase 
are  unknown.  Bach  ^"^'^  obtained  evidence  that  tyrosinase  is  not  a 
specific  oxidizing  enzyme,  but  consists  of  an  aminoacidase,  which  dis- 
integrates the  tyrosine  and  makes  it  susceptible  to  the  action  of  pheno- 
lase  which  is  the  oxidizing  agent,  v.  Fiirth  and  Schneider  found  the 
product  of  oxidation  of  tyrosine  by  animal  tyrosinase  related  to  cer- 
tain of  the  melanins  of  animal  tissues,  and  believe  that  tyrosinase  is 
responsible  for  the  production  of  many  normal  pigments.  In  the 
ink-sacs  of  the  squid,  which  eject  an  inky  fluid  containing  melanin- 
like pig-ment,  tyrosinase  was  also  found,  corroborating  this  hypothesis, 
and  it  is  probable  that  tyrosinase  in  the  skins  of  aninmls  is  responsible 
for  their  pigmentation.*"  Bacteria  also  contain  tyrosinase,""  and  this 
or  similar  enzymes  seem  to  be  present  in  melano-sarcomas."^ 

Gonnermann "-  found  that  tyrosinase  from  beet-root  produced 
homogentisic  acid  by  acting  on  tyrosine,  which  is  of  interest  in  con- 
nection with  the  congenital  hereditary  disease,  alkaptonuria  {q.  v.)y 
in  which  the  urine  becomes  dark  upon  exposure  because  of  the  pres- 
ence of  homogentisic  acid.  The  action  of  tyrosinase  upon  the  aromatic 
radicals  of  proteins  is  of  great  importance  in  the  study  of  both 
physiological  and  pathological  pigment  formation,  and  hence  has  re- 

ssBattelli  and  Stern.   Biocliem.   Zeit.,   1910    (29),   130. 
ssaBiochem.  Zeit.,  1914    (60).  221. 

89  Meirovvsky,  Cent.  f.  Path.,  1909   (20),  301. 

90  Lehmann  "and  Sano,  Arch.  f.  Hyg.,  1908    (67),  99. 

91  Alsberg,  Jour.  Med.  Res.,  1907' (16).  117:  Neuberfj:,  Virchow's  Arcliiv.,  190S 
(192),  514;  Gessard,  Compt.  Rend.  Soc.  Biol.,  1902   (54),  1305. 

92  Pfluger's  Arch.,  1900    (82),  289. 


74  ENZYMES 

ceived  extensive  study,  which  will  he  found  fully  described  in  the 
monograph  b}'  Kastle  {loc.  cit.)  '^*  and  under  the  appropriate  subjects 
in  subsequent  chapters. 

Other  Oxidizing  Enzymes. — Of  the  great  numlier  of  other  less 
studied  oxidizing  enzj-mes  little  can  be  definitely  stated.  Some  con- 
sider that  they  are  largely  different  manifestations  of  the  action  of 
one  oxidizing  ferment,  but  against  this  view  Jacoby  mentions  that  they 
occur  distributed  unequally  in  different  organs,  can  be  separated  from 
each  other,  and  they  cause  different  reactions.  For  the  catalase  and 
for  laccase  (which  produces  the  Japanese  lacquer  by  an  oxidizing 
process)  and  perhaps  for  other  oxidizing  ferments,  iron  and  man- 
ganese may  be  essential  constituents.  Of  particular  significance  for 
pathology  are  the  enzymes  which  accomplish  the  oxidation  of  purines 
to  uric  acid  and  the  subsequent  destruction  of  uric  acid.  These  are 
discussed  in  Chapter  xxi.  Also  the  enzymatic  oxidation  and  reduc- 
tion of  ^-oxj^butyric  acid  and  aceto-acetic  acid  in  the  liver,  as  studied 
by  Dakin  and  Wakeman,"^  are  of  great  importance  in  acidosis  {q.  v.). 

Reducing  enzymes  have  not  yet  been  satisfactorily  demonstrated."* 
It  is  possil)le  that  they  do  not  exist,  and  that  the  intracellular  re- 
ductions that  are  carried  on  within  the  cells  are  brought  about  by 
simple  chemical  reactions  independent  of  catalysis.  The  best  known 
intracellular  reduction  is  that  of  methylene  blue,  which  can  be  readily 
studied  experimentally  because  the  blue  color  disappears  on  reduction 
of  the  dye.  It  is  open  to  question  if  this  particular  reduction  is  due 
to  a  reducing  enzyme.  According  to  Ricketts  ^^  the  reduction  depends 
upon  two  bodies,  one  thermostabile,  the  other  thermolabile,  recalling 
the  reaction  of  complement  and  amboceptor.  Strassner  ^"^  found  evi- 
dence that  the  SH  groups  of  the  tissues  are  responsible  for  the  reduc- 
tion of  methylene  blue ;  their  activity  is  impaired  by  heating,  but  a 
thermostable  element  of  tissues  augments  the  reducing  activity  of  SH 
compounds,  thus  corroborating  and  explaining  the  observations  of 
llicketts.  Harris,"^  however,  believes  that  the  evidence  for  the  exist- 
ence of  a  true  reducing  enzyme  is  as  good  as  for  most  other  cellular 
enzymes.  An  enzyme  has  been  found  in  the  liver,  muscle  and  kidney 
which  transforms  aceto-acetic  acid  into  l-/3-oxybutyric  acid,  and  called 
ketoroductase   (Friedmann  and  Maase)."'^" 

Oxidizing  Enzymes  in  Pathological  Processes. — Although  the 
oxidizing  enzymes  undoubtedly  play  an  important  part  in  pathologi- 
cal conditions,  they  have  been  but  little  investigated  from  this  stand- 
point. Jacoby  found  that  they  did  not  disappear  from  the  degen- 
erated liver  in  phosphorus  poisoning  or  in  diabetes,  or  when  the  liver 

03  Jour.   .Amor.  IVfod.  Assoc,   1910    (54),   14-11. 

o4Soo  TIofTtPr.  Arc-li.  exp.  Path.  u.  Pharm..  lOOS,  Suj)])!..  p.  25.3. 

"•"•Jour  of  Infoft.ions   Disoaaos,   1004    (1),   500. 

(•«  Biocliem.   Zcit.,    1010    (20),  205. 

"7  Biofiioni.  .loiir.,   1010    (5),   14:?. 

07a  iJiocliom.  Zeit.,   1012    (27),  474;    101.3    (55),  4,58. 


(}\inr/.i\(!  i:\'/.y\ii:H  75 

undergoes  self-digestion,  which  speaks  against  Sjjitzer's  contention  that 
oxidase  is  a  nucleoprotein.'-''*  JSelilesinger '-''''  found  that  it  is  less  in 
amount  in  livers  of  children  dead  from  gastro-iutestinal  diseases  than 
in  normal  livers,  as  also  did  Briining.^  I  am  inclined  to  believe  that 
fatty  metamorphosis,  wlien  brought  about  by  poisons,  is  often  due 
to  inhibition  of  the  oxidizing  enzymes  (v.  fatty  metamorphosis), 
although  I  found  that  livers  the  seat  of  the  most  profound  fatty  de- 
generation showed  no  evident  impairment  of  their  power  to  oxidize 
xanthine  and  uric  acid.-  Buxton^  failed  to  find  in  tumors  any  en- 
zyme giving  the  guaiac  test  alone,  but  found  enzymes  that  did  so  in 
the  presence  of  HgOg  (peroxidases).  Catalase  was  present,  but  no 
very  positive  reactions  for  oxidizing  enzymes  were  obtained  by  the 
indo-phenol  reaction,  the  hydrochiuon  reaction,  or  with  tyrosine  for 
tyrosianase.  v.  Fiirth  and  Jerusalem  *  have  found  evidence  tliat  the 
melanin  of  melanotic  tumors  of  horses  is  produced  by  tyrosinase. 
Peroxidase  has  been  demonstrated  in  the  granules  of  pus  cells 
iFisclieP). 

Meyer  ®  found  that  leucocytes,  whether  from  pus  or  leukemic  or 
pneumonic  blood,  contained  a  substance  oxidizing  guaiac  directly, 
without  the  presence  of  H^O,,  which  is  not  liberated  until  the  cells 
are  destroyed.  By  microchemical  reactions  oxidases  have  been  found 
present  in  the  myelocytes  and  nucleated  erythrocj'tes  in  leukemia,  be- 
ing absent  from  the  polynuclear  cells.'  The  observation  of  Natalie 
Sieber  *  that  oxidases  of  the  blood  and  of  vegetable  origin  destroy 
diphtheria  toxin  rapidly,  and  also  tetanus  toxin  and  ricin,  has  been 
confirmed  by  Loewenstein  as  far  as  destruction  by  peroxide,  with  or 
without  the  presence  of  catalase,  is  concerned.  Oxidation  is  un- 
doubtedly an  important  process  in  defending  the  bod}'  against  other 
forms  of  poisons,  including  the  so-called  ' '  fatigue  toxins, ' '  and  Battelli 
and  Stern  consider  that  all  the  oxidizing  enzymes  so  far  definitely 
identified  are  concerned  only  in  protective  processes.  (See  Chapter 
vii).  Schmidt"  has  found  that  by  oxidation  certain  poisonous  mor- 
phin  derivatives  are  rendered  non-poisonous  by  liver  extracts.  Oxalic 
acid  and  poisonous  fatty  acids  are  also  oxidized  into  harmless  sub- 
stances; phosphorus  and  sulphur  are  oxidized  into  their  acids,  which 

9s  Duccheschi  and  Almagia  (Arch.  ital.  Biol.,  1903  (39),  29)  also  found  tlie 
aldeliydase  in  livers  of  phosphorus  poisoning  usually  no  less  abundant  tlian  in 
normal   livers. 

9'J  Hofmeister's  Beitr.,  1903  (4),  87. 

iMonat.  f.  Kinderheilk.,   1903    (2),   129. 

2. Tour.  Exper.  Med.,  1910   (12),  607. 

3. Jour.  Med.  Research,  1903  (9),  356. 

4  Hofmeister's  Beitr.,  1907    (10),  131. 

5\Vien.  klin.  Woch.,  1910   (23),  1557. 

sMiinch,    med.   Woch.,    1903     (50),    1489. 

7  Fiessinger  and   Roudowska,   Arch,   de   med.   exper.,    1912    (24),   585. 

sZeit.  physiol.  Chem.,  1901    (32),  573. 

9  Dissertation,  Heidelberg,   1901. 


76  ENZYMES 

are  then  neutralized.     Indole  and  skatole  are  oxidized  into  less  harm- 
ful substances. 

The  Indophenol  Reaction. i" — Alplui-naplitliol  and  dimethyl-para-plicnylendia- 
min.  wlieii  lirouglit  louctln'r  in  alkaline  solntioii,  become  oxidized  in  the  pres^ence 
of  air  and  form  an  insoluble  blue  dye,  indophenol.  Tliis  reaction  is  jireatiy 
accelerated  by  oxidizing'  agents,  and  it  has  liien  found  that  certain  tissues  pos- 
sess this  property,  hence  the  indophenol  synthesis  has  been  used  for  microchemical 
study  of  the  presence  and  distribution  of  oxidizing  enzymes  in  cells.  As  the  in- 
tracellular agent  which  causes  this  reaction  is,  ho\.ever,  so  resistant  to  heat  and 
chemicals  that  it  can  be  demonstrated  in  sections  fixed  in  formalin  and  prejiared 
by  the  ordinary  paralbn  imbedding  method  (Dunn),  there  is  room  for  much  doubt 
as  to  whetlier'  it  represents  a  true  enzyme  of  the  polyphenol  oxidase  class.  It 
may  be  that  it  is  identical  with  phenolase.n  In  tiie  presence  of  small  amounts 
of  peroxide  tiie  granules  of  leucocytes  and  myelocytes  are  stained  with  alpha- 
naphthol  alone,  which  Graham  na  interprets  as  oxidation  by  an  enzyme  of  the 
peroxidase  type.  The  indophenol  reaction  is  observed  best  in  the  granules  of 
neutroi>hile  leucocytes  of  blood  and  in  myeloid  cells  of  bone  marrow,  leukemic 
blood  and  fetal  organs;  eosinophiles  and  basophile  leucocytes  also  give  reactions, 
but  not  lymphocytes,  mature  erythrocytes,  or  most  fixt  tissue  cells.  ( See  Dunn, 
Quart.  Jour.  Med.,  1913  (6),  293.)  By  using  alkali-free,  unfixt  tissues  Gierke 
found  granules  present  in  tissue  ceils  generally,  and  Griiff  states  that  they  occur 
in  proportion  to  the  metabolic  activity  of  the  cells;  they  are  abundant  in 
carcinomas,  scanty  in  sarcoma  and  connective  tissue  growths  generally,  arc  not 
destroyed  in  cloudy  swelling  or  fatty  changes,  but  disappear  in  infarcts  and 
autolyzing  tissues,  and  in  tissues  asphyxiated  with  illuminating  gas.i-  Lung 
tissue  is  especially  poor  in  this  form  of  oxidative  activity,i2a  but  giant  cells  of 
tubercles  contain  oxidase  granules. 12b  During  experimental  pneumococcus  sep- 
ticemia the  indophenol  oxidase  reaction  is  decreased  in  the  tissues. 12c 

Glycolytic  Enzymes.^^ — The  oxidation  of  sugar  by  the  tissues, 
which  is  one  of  the  chief  sources  of  energy  in  the  animal  body,  presum- 
ably takes  place  through  several  steps.  Of  these,  it  is  believed  by  some 
that  the  first  is  the  formation  of  glycuronic  acid — 


CH,OH— (CHOH),C— H  +   0,  =  COOH— ( CHOH )  ,C— H  +  H,0, 
(glucose)  (glycuronic  acid) 

but  the  subsequent  changes  which  involve  decomposition  of  the 
straight  chain  are  not  at  present  understood.  Attempts  to  isolate 
from  various  organs  an  enzyme  oxidizing  glucose,  particularly  from 
the  pancreas,  muscle,  and  liver,  have  led  to  varying  results  and  nnich 
dissension,  but  it  is  probable,  because  of  these  failures,  that  no  such 
enzyme  exists  in  quantities  sufficient  to  account  for  the  amount  of 

10  Literature  given  bv  Schultze,  Ziegler's  Beitr.,  1909  (45),  127:  Dunn.  Jour. 
Path,  and  Bact.,  1910   ('l5),  20;  Griiff,  Frankfurter  Zeit.  f.  Path.,  1912.(12),  358. 

11  Bach  and  Marvanovitsch,  Biochem.  Zeit..  1912    (42),  417. 
1  la  Jour.   Med.    Ues.,  .1916    (35),   231. 

12  See  Klojifcr,  Zeit.  exp.  Pharm.,  1912    (11),  407. 
12a  Weiss,  Wien.  klin.  Woch.,  1912   (25),  t)97. 
i2bMakino,  Verh.  Jai)an.   Patli.  Gesell.,   1915    (5),  71. 
i2.Medigrcceanu.  Jour.  Exp.  .Med.,  1914    (19),  ,303. 

13  Also  discussed  under  "Diabetes,"  chap.  xxil.  As  glycolysis  by  blood  and 
tissues  can  <)(<nr  without  oxygen,  Battelli  and  Stern  exclude  the  glycolytic  from 
the  oxidizing  enzymes. 


Lii'Asi-:  77 

sugar  coinbustiou  that  is  iionnally  a('r()in|)lished.  0.  Coliiiheim  '^^  at- 
toinjited  to  explain  the  failures  by  his  observation  that  the  panereas 
])roduees  a  substanee  that  aetivates  an  inactive  glycolytic  en/ynie  in 
the  muscles,  liver,  and  probably  in  other  organs.  This  work  has  been 
much  contested,  and  is  not  generally  accepted,  so  we  are  still  in  the 
dark  as  to  how  the  carbohydrate  oxidations  are  accomplished.  (See 
Cliai)ter  xxii.) 

LIPASE  n 

Lipase  is  probably  present  in  greater  or  less  amount  in  ail  cells. 
In  the  discussion  of  the  reversible  action  of  enzymes,  on  a  previous 
page,  the  modern  conception  of  fat  metabolism  has  been  exi)lained, 
which  considers  it  to  depend  upon  the  existence  of  lipase  in  the  cells 
and  fluids  throughout  the  body.  On  account  of  the  technical  diffi- 
culties in  the  w^ay  of  using  higher  fats,  such  as  triolein,  in  experi- 
2uental  work,  the  esters  of  lower  fatty  acids  have  generally  been  used, 
particularly  ethyl  butyrate,  salicylic  acid  esters,  and  glycerol  triace- 
tate. Enzymes  splitting  ethyl  butyrate,  and  presumably  higher  fats, 
have  been  demonstrated  in  practically  all  tissues  examined ;  the  names 
of  Kastle  and  Loevenhart  in  this  country,  and  Hanriot  in  France, 
being  particularly  connected  with  this  work.  Whether  in  all  cases 
the  presence  of  this  reaction  is  proof  positive  of  the  presence  of  an 
enzyme  splitting  fats,  a  true  "lipase,"  is  not  yet  known.  JMuch  of 
the  work  so  far  reported  on  the  occurrence  of  lipase  in  tissues  is  of 
questionable  value,  especially  as  to  quantitative  results,  because  of 
faulty  methods.  Saxl  ^■"'  points  out  and  avoids  some  of  these  errors, 
and  finds  that  during  autolysis  of  tissues  the  splitting  of  the  natural 
fats  present  in  the  cells  is  but  slight ;  simple  esters  are  attacked  more, 
especially  amyl-salicylate ;  muscle  and  blood  are  the  least  active 
tissues.  Most  authors  agree  that  lymphoid  cells  are  especially  rich  in 
lipolytic  enzymes.^'  In  the  serum  of  normal  individuals  the  esterase 
content  seems  to  be  quite  constant,^"'^  and  Quinan  ^-'^  found  the  tissue 
content  also  constant,  the  liver  containing  about  twice  as  much  as 
the  kidney  and  over  three  times  as  much  as  the  muscle.  He  states 
that  different  parts  of  the  brain  have  characteristic  lipase  activity 
(but^-rase).^'"'  Thiele  ^"'^  has  found  that  blood,  chyle,  and  various 
tissues  also  contain  an  enzyme  which  can  hydi'olyze  lecithin,  but  except 

isaZeit.  phvsiol.  Cliem.,  1903  (39),  330;  also  see  Simpson,  Bioclicin.  .l..ur..  1910 
(5),    126. 

i-t  For  literature  on  lipase  see  Connstein,  Ergebnisso  Pliysiol.,  1904  (3.  Aht.  1), 
194;  concerning  the  behavior  of  lipase  see  Taylor,  Jour.  Biol.  Cheni.,  1906  (2), 
103:   Fali<,  Proc.  Natl.  Acad.,  1915    (1),   ]3(i. 

isBiochem.  Zeit.,   1908    (12),  343. 

1"  The  distribution  of  lipases  in  difTerent  species  of  animals  and  their  various 
organs  has  been  investigated  bv  Porter,  Miinch.  med.  Woch.,  1914   (61).  1774. 

isaSagal,  Jour.  Med.  Pes.,  1916  (34),  231. 

^shJMd.,  191.5    (32),  45. 

15c  Ibid.,  1916    (35),  79. 

isdBiocliem.  Jour.,  1913   (7),  275. 


78  ENZYMES 

in  the  pancreas  it  does  not  hydrolyze  neutral  fats.  The  brain  contains 
enzymes  ]iydrolyzin<>-  mono-  and  triaeetin,  lecithin  and  cephalin.^^^ 

Little  is  known  about  the  part  played  by  lipase  in  pathological  con- 
ditions. According  to  Achard  and  Clerc/**  the  amount  of  splitting  of 
ethyl  butyrate  by  the  blood-serum  is  lessened  in  most  diseases,  and  in- 
creases and  decreases  Avith  the  health  of  the  patient;  according  to 
Pribram  ^*  and  Sagal  ^''^  it  is  increased  in  the  blood  during  fevers. 
Clerc  ^'•'  found  that  acute  arsenic,  phosphorus  and  diphtheria-toxin 
poisoning  increased  this  property  of  the  serum,  while  chronic  poison- 
ing and  staphylococcus  intoxication  lowered  it.  Somewhat  similar 
results  were  obtained  by  Grossmann,-"  but  Saxl  found  no  increased 
activity  in  phosphorus  poisoning.  Using  the  ethyl  butyrate  test, 
AYintemitz  and  ]Meloy  -^  found  that  the  more  nearly  normal  an  organ 
is  the  more  cleavage  of  the  ester;  lipolytic  activity  is  low  at  birth, 
increases  rapidly  during  the  first  few  days  of  life,  and  does  not  de- 
crease in  old  age.  There  is  a  decline  in  activity  of  tissues  in  diabetes, 
tuberculosis,  and  the  toxemia  of  pregnancy,  in  the  livers  of  passive 
congestion  and  fatty  degeneration,  in  the  pneumonic  lung  and  the 
cirrhotic  liver.  After  taking  food  there  is  a  slight  increase  in  esterase, 
reaching  a  maximum  in  three  hours. -'''^  Wliipple  -'^°-  finds  the  blood 
lipase  (butyrase)  increased  whenever  there  is  injury  to  the  liver, 
such  as  in  chloroform  anesthesia  and  puerperal  eclampsia ;  it  is  lowered 
in  cirrhosis.  Poulain  --  found  that  the  butyric-splitting  power  of 
lym])h-g]ands  draining  infected  areas  Avas  decreased.  Fisclier  ^^ 
observed,  in  a  case  of  extreme  lipemia  in  diabetes,  that  the  lipolytic 
power  of  the  blood  was  absent.  The  lipase  of  lipomas  presents 
no  demonstrable  difference  from  that  of  ordinary  fatty  areolar 
tissues.-* 

Lipase  has  also  been  demonstrated  in  pus  by  a  number  of  ob- 
servers,-' Avho  agree  that  there  is  more  in  exudates  than  in  transu- 
dates. Zeri  -"  found  lipase  in  the  urine  only  when  pus  or  blood  was 
also  present,  but  Pribram  and  Loewy  -'^  found  it  in  nephritis,  con- 

isoEnplisli  and  IVIacArtlnir  (Jour.  Aiiicr.  Chom.  Soc.  1915  (.37).  Or).'?).  who 
have  also  found  in  sheop  l)rain,  erepsin,  amylase,  catalase.  enzymes  deeomposinfj 
arluitin  and  salol,  probably  pepsin  and  trypsin,  but  not  peroxidase,  oxidase, 
reductase,  jjuanase,  urease  or  rennin. 

i«  Compt.  Rend.  Soc.  Biol..  1002   (54),  1144. 

IS  Cent.  inn.  Med.,  1908    (29),  81. 

loCompt.   Rend.   Soc.  Biol.,    1901    (53),    1131. 

zoBiochem,  Zeit.,  1912    (41),  181. 

21  Jour.  Med.  Res.,  1910  (22),  107. 
20a.Toblinfr  et  al..  Jour.  Exp.  Med.,  1915   (22),  129. 

2ia  Whipple  et  al.,  Bull.  Jolms  Hopkins  Hosp.,  1913   (24),  207  and  357. 

22  Com  p.  Rend.  Soc.  Biol.,  1901    (53),  7Sfl. 
23Virchow's  Arch.,   1903    (172),  218. 

24  Wells,  Arch.  Int.  ]\led.,   1912    (10),  297. 

2'-.  Aclialme.  C'ompt.  Rend.  Soc.  Biol.,  1899    (51),  5(iS -.    Zeri.  11  Bolicliiiieo.  1903 
(10),  43.-];   Meiiniii,  Clin.  ]\led.  Ital.,  1905   (44),  129. 
2"  11   Policlinico.   1905    (12),   733. 
27  Zeit.   j)l)ysiol.  f'liein.,   1912    (70),  489. 


LIPASE  79 

gestion,  polyuria  and  other  conditions.  Lorenzini,-'''^  however,  re- 
ports that  in  albuniinnria  tlie  lipase  content  of  the  urine  is  reduced, 
in  common  with  other  enzj-mes,  there  being  a  simultaneous  accumula- 
tion of  enzymes  in  the  blood. 

Fiessinger  and  ]\Iarie  -*  contend  that  the  lymphocytes  of  exudates 
are  the  chief  source  of  lipase,  and  sug:gest  that  this  may  be  of  effect 
in  defense  against  the  fatty  tubercle  bacilli.  Toxines  were  found  by 
Pesci  -^  to  increase  the  butyrase  but  not  the  other  lipases  of  liver 
tissue.  In  syphilis  the  lipolytic  activity  of  the  serum  is  increased,^" 
which  may  be  related  to  Bergell's^^  observation  on  the  origin  of 
lipase  in  lymphocytes  (corroborating  Fiessinger  and  ^Marie).  Jobling 
and  Bull  ^-  state  that  a  specific  serum  lipase  increase  occurs  in  animals 
immunized  to  red  corpuscles,  and  that  this  lipase  has  to  do  with 
hemolysis ;  but  ^Mendel  ^^  found  no  evidence  that  hemoh^sis  by  ricin  is 
related  to  lipase.  Abderhalden  and  Rona  ^*  found  that  excess  feeding 
of  fats  leads  to  an  increase  in  the  lipase  of  the  blood. 

The  part  played  by  lipase  in  fatty  degeneration  must  be  of  great 
importance,  but  as  yet  it  has  been  little  considered,  except  that  Loeven- 
hart.  and  Duccheschi  and  Almagia  '"'  found  no  appreciable  difference 
in  the  lipase  content  of  nonnal  and  phosphorus-poisoned  livers,  but 
in  chloroform  poisoning  Quinan  ^^'^  found  a  decrease  in  the  butyrase 
of  the  liver,  although  it  was  increased  in  the  kidneys  and  muscles. 
This  question  will  be  considered  more  fully  in  discussing  fatty  meta- 
morphosis. 

An  improved  method  of  testing  for  lipase  action  has  been  devised 
by  Rona  and  Michaelis,^®  by  measuring  the  change  in  surface  tension 
caused  by  hydrolysis  of  a  soluble  ester,  usually  tributyrin.  Using  this, 
Bauer  found  that  every  human  serum  contains  fat-splitting  enzymes, 
which  are  greatly  decreased  in  carcinoma  and  advanced  phthisis,  some- 
what decreased  in  syphilis  and  exophthalmic  goitre,  and  increased  in 
early  pulmonary  tuberculosis.  Caro  ^'^^  found  a  decrease  in  all  cases 
of  cachexia,  but  there  was  no  relation  between  the  lipolytic  enzyme 
and  the  blood  picture.  The  blood  contains  no  thermostable  antilipase 
analogous  to  the  antitrypsin.  Red  corpuscles  are  said  to  contain  an 
enz\'me  splitting  cholesterol  esters,  " cholesterase."  ^'^ 

27a  Policlinico,   1015    (22),  35S. 

2sCompt.  Rend.  Soc.  Biol.,  1909  (67),  177.  See  also  Eesch.  Dent.  Arch.  klin. 
Med.,  1915    (118),  179. 

29  Pathologrica.  1912   (.3),  207. 

30  Citron  and  Reieher,  Berl.  klin.  Woch.,  1908    (45),  1.39S. 

31  Miinch.  med.  Woch.,  1909    (56),  64. 

32  .Jour.  Exp.  Med..  1912    (16),  48.3. 

33  Arch.  Fisiol.,  1909   (7).  168. 
34Zeit.  phvsiol.  Chem..  1911    (75).  30. 

35  Arch.  Ital.  Biol.,  1903    (39).  29. 
35a  .Jour.  :\Ied.  Res.,  1915    (32),  73. 

36  See  Bauer,  Wien.  klin.  Woch..  1912   (25),  1376   (bibliography). 
3Ga  Zeit.  klin.  Med.,  1913    (781,286. 

37  See  Cytronberg,  Biochem.  Zeit.,  1912   (45),  281. 


80  i:\'/.)MEH 

Fat  necrosis,  resulting  from  the  escape  of  pancreatic  juice  into  the 
peripanereatic  tissues  and  abdominal  cavity,  undoubtedly  is  largely 
the  result  of  lipase  action.  (See  "Fat  Necrosis,"  Chapter  xiii,  for 
complete  consideration.) 

AMYLASE  3H 

Although  undt'i*  ordinary  conditions  starch  is  not  supposed  to  enter 
the  blood  stream  and  tissues,  yet  all  tissues  and  body  fluids  are  capable 
of  hydrolyzing  starch.  Apparently  the  amylase  is  derived  from  the 
pancreas  and  salivary  glands,  and  i)ossibly  from  many  or  all  other 
tissues  (King),  but  it  is  not  (piantitatively  related  to  the  amount  of 
carbohydrate  in  the  diet  of  a  species  or  an  individual  (Carlson  and 
Luckhardt).  In  llic  bloiMJ  it  occurs  in  the  albumin  fraction.'"'  There 
is  disagreement  in  the  literature  as  to  the  variations  in  amount  of 
amylase  in  the  blood  during  disease,  and  little  information  concerning 
its  distribution  in  the  tissues.  Normally  the  kidneys  and  liver  seem 
to  be  most  active.  During  acute  infections  the  blood  amylase  is  in- 
creased, presumably  coming  from  the  leucocytes  (King).  It  is  greatly 
increased  when  the  pancreas  is  acutely  inflamed  or  injured  (Stocks). 
Intravenous  or  subcutaneous  injection  of  starch  is  said  to  increase  the 
blood  amylase,  presumably  as  a  defensive  reaction  (Abderhalden), 
but  the  amylase  ordinarily  in  the  blood  seems  to  be  a  waste  substance 
on  its  way  to  excretion,  rather  than  a  functionating  enz^one  of  the 
blood.     There  appears  to  be  no  normal  antiamylase  in  the  blood. 

Because  of  possible  diagnostic  signiticance,  the  amylolytic  activity 
of  the  urine  has  been  particularly  studied,  and  found  normally  to  be 
approximately  constant  for  24  hour  specimens  of  the  same  individual.*" 
Anything  impairing  the  excretory  capacity  of  the  kidney  decreases  the 
urinary  amylase,  although  sometimes  when  the  urine  contains  blood, 
pus,  or  much  all)umen  there  may  be  an  increased  amylase  excretion  in 
spite  of  diminished  functional  activity.  There  may  be  an  increase  in 
the  amylase  in  the  blood  when  the  urinary  amylase  is  decreased,  but 
with  normal  kidneys  increase  of  the  blood  amylase  causes  an  increase 
in  the  urine;  hence,  acute  pancreatic  diseases  cause  an  increased 
urinary  amylase  TStocks),  but  this  is  not  constant  (McClure  and 
Pratt).  In  diabetic  urine  it  is  said  to  l)e  usually  decreased,  but  this 
is  mostly  accounted  for  by  the  dilution  of  the  urine.  Parenteral  in- 
jection of  stai-ch  causes  a  marked  increase  in  the  amount  of  diastase  in 
the  urine   (King)  .'^^ 

3«  Litcraluro  {.nvcn  bv  Kinff,  .Amor.  Jour.  Phvsiol..  1914  (S.'i),  301;  Govelin, 
Arc-h.  Int.  Med.,  1014  (13),  06;  Storks.  Quart.  .Toiir.  ^rcd.,  1010  (0),  210;  :^[c'Clure 
and  Pratt.  An-h.  Int.  Med.,  1017  HO).  .'>GS. 

3»Satta,  Arcli.  Sci.  "MM.,  lOlo    (.10),  4(i. 

■">  In  infants  llio  urino  anivlaso  is  low  f^[c("'Iuro  and  riiaiiccUdr,  Zoit.  Kindor- 
hcilk..   1014    (11),  4S:{. 

'I  rrnr.    So.-.    Kvii,    lii.il,.    I'.HT     (  1.-)).    1(11. 


CHAPTER    III 

ENZYMES  (Continued) 

INTRACELLULAR   PROTEASES'    (PROTEOLYTIC    ENZYMES),    IN- 
CLUDING A  CONSIDERATION  OF  AUTOLYSIS 

To  what  extent  synthesis  of  proteins  goes  on  in  the  body  is  still  a 
problem ;  still  more  uncertain  is  the  part  played  by  reversible  action 
of  proteases.  There  is  evidence  enough  that  somewhere  in  the  body 
the  amino-acids  can  be  rebuilt  into  protein,  for  several  investigators 
have  succeeded  in  keeping  animals  in  nitrogenous  equilibrium  by  feed- 
ing them  products  of  proteolysis  that  contained  no  protei-us  whatever, 
and  as  the  proteins  of  the  animal  body  are  being  broken  down  in- 
cessantly, it  must  be  that  they  were  replaced  by  synthesis  of  the  non- 
protein material  fed  to  the  animals.  In  addition,  it  has  long  been 
questioned  whether  amino-acids  absorbed  from  the  intestines  are  not 
resynthesized  into  proteins  while  passing  through  the  intestinal  wall. 
Cohnheim  found  that  in  the  intestinal  epithelium  there  is  an  enzyme, 
erepsin,  capable  of  splitting  albumoses  and  peptones  into  the  amino- 
acids,  which  enzyme  presumably  exists  for  the  purpose  of  securing 
complete  cleavage  of  all  ingested  proteins  into  their  ultimate  "build- 
ing stones."  This  may  be  looked  upon  as  a  provision  to  reduce  all 
varieties  of  proteins  to  their  common  elements,  so  that  the  body  by 
quantitative  selection  can  resynthesize  them  into  its  own  types  of 
protein,  for,  as  is  well  known,  foreign  proteins  (e.  g.,  egg-albumin) 
introduced  directly  into  the  blood  stream  cannot  be  utilized,  but  are 
excreted  unaltered  in  the  urine. ^  As  was  shown  for  lipase,  the  as- 
sumption that  such  synthesis  occurs  as  a  normal  physiological  process 
by  reverse  enzyme  action,  requires  that  the  proper  enzymes  be  present 
in  the  cells  throughout  the  body,  and  within  the  past  few  years  it  has 
been  abundantly  demonstrated  that  such  is  the  case. 

For  over  half  a  century  it  has  been  known  that  amebre  digest  solid 
proteins  within  their  bodies,  but  it  is  only  within  a  few  years  that 
proteolytic  enzymes  have  been  definitely  isolated  from  them.  It  has 
been  nuieh  the  same  w'ith  the  intracellular  proteases  of  the  higher 

1  As  the  possibility  exists  tliat  ferments  T\-hi<ii  digest  proteins  may  lie  able  to 
perform  a  certain  amount  of  synthesis  of  proteins,  the  term  "proteolytic  enzyme" 
seems  to  be  less  suitable  than  the  term  "iirotease,"  which  merely  means  an  enzyme 
acting  on  proteins,  and  does  not  compel  us  to  accept  any  particular  view  as  to 
what  the  action  is. 

2  According-  to  Austin  and  Eisenbrey  (Arch  Int.  Med..  1912  (10),  ,305),  dogs 
on  a  nitrogen-free  diet  can  utilize  horse  serum  injected  intravenously. 

6  81 


82  EXZYMEfi 

organisms.  In  1871  Iloppe-Seyler  referred  to  the  liquefaction  of 
dead  tissues  within  the  body  which  occurred  without  putrefaction, 
and,  as  he  noted,  resembled  the  effects  of  the  digestive  ferments.  It 
was  nearly  twenty  years  later  that  Salkowski  •'  showed  definitely  that 
tliis  softening  of  dead  tissues  was  really  brou^lit  al)out  through  a  true 
digestion  by  intracellular  enzymes,  which  produced  the  same  splitting 
products  that  were  at  that  time  considered  characteristic  for  tryptic 
digestion  (leucine  aiui  tyrosine).  'I' lie  process  he  named  "autodiges- 
twn."  This  important  observation  i-enuiined  almost  unnoticed  for 
ten  years  more,  when  Jacoby,'  in  1900,  took  up  the  investigation  of 
this  matter  of  cellular  self-digestion,  and  after  this  the  importance  of 
the  principles  involved  became  for  the  first  time  generally  appreciated. 
Jacoby  rechristened  the  process  ''autolysis,"  by  which  name  it  is  now 
commonlv  known. 

AUTOLYSIS ' 

Autolysis  is  generally  studied  by  the  nictliod  used  by  Salkowski, 
which  depends  upon  the  difference  in  the  susceptibility  of  bacteria 
and  of  enzymes  to  antiseptics.  The  organs  are  ground  to  a  pulp, 
placed  in  flasks  with  or  without  the  addition  of  water  or  dilute  acids, 
and  bacterial  action  is  prevented  by  the  addition  of  antiseptics  that 
are  not  i)oisonous  to  enzymes — toluene  and  chloroform  are  most  com- 
monly used.  It  is  possible  also  to  secure  organs  in  an  aseptic  con- 
dition and  to  permit  them  to  undergo  autolysis  without  the  use  of 
antiseptics,  but  the  practical  difficulties  are  such  lliat  this  method 
is  seldom  used — it  is  sometinu's  desigiuited  as  "aseptic  autohisis,"  in 
contradistinction  to  antiseptic  autolysis  by  the  Salkowski  method.  In 
a  short  time  it  can  be  seen  that  digestive  changes  have  taken  place, 
particularly  if  comparisons  are  made  with  control  flasks  in  which 
the  enzymes  have  been  destroyed  by  boiling.  To  determine  the  rate 
of  autolysis  the  amount  of  nitrogen  that  renuiins  in  the  form  of 
coagulable  compound.s,  and  that  which  is  converted  into  soluble,  non- 
coagulable  compounds  (albumoses,  peptones,  annnonia  compounds, 
amino-acids,  etc.),  is  compared.  The  method  nuiy  be  illustrated  by  a 
concrete  e.xamjjle :  A  given  specimen  of  enndsioni/.cd  liver  tissue  was 
permitted  to  digest  itself  for  twenty -two  days.  At  the  end  of  that 
time  39.4  per  cent,  of  the  nitrogen  was  still  contained  in  the  com- 
pounds that  remained  insoluble  or  became  so  after  the  autolysis  was 
stojjped  by  boiling;  while  60.6  ])vv  cent,  of  the  nitrogen  was  in  a 
soluble  form.  A  control  S]iecini('n  from  the  same  liver  Avas  boiled 
while   fresh    to   kill    the   eii/.ynies.   ;in<l    then    let    stiind    under   the   same, 

■i  Zcit.  f.  kliri.  Med.,  1890,  supi.loim-iit   In  I'.d.   17.  p.  77. 

<  Zcit.  f.  pli.vsiol.  Clipni.,  1!K)()    (3(l|.    MM. 

•'•  Hr'sinnC'  of  litfriitiirc  Iiy  Salkowski.  DcuLsi-lie  Klinik.  1003   (11),  147;  also  see- 
Sclilesiiiticr.   llofiiu'is<cM's    licit  rii-zc.   l!Mt:{    (4).   87:    Oswald.    Uioclicin.   Coiitr..    1005 
CM.    'Ml')-.    I.cvciic.   .lour.   AiiKM-.    Med.    .\sso<'..    ]'MM\    MC.i,    77(1.      Sec   also    \"i  •ollc. 
Ann.  Inwl.   I'aslcur,  101.3    (27),  07:   von   ImhIIi.  ••(li.ini'^t  r\    of   Mctaholisni.""    \inor 
TruMHl.,   10i«. 


MTOfASIS  83 

conditions.  Tn  tliis  specimen  1)0.4  per  cent,  of  the  nitrogen  was  in 
an  insoluble  form,  and  9.6  per  cent,  was  soluble.  Therefore,  over 
half  of  all  the  protein  of  the  liver  had  been  changed  into  non- 
coagulable  substances  in  the  course  of  about  three  weeks  (at  37°  C). 
Complete  disintejiration  of  the  proteins  with  liberation  of  all  the 
amino-acid  complexes  is  probably  never  readied.  Of  45.8  grams  of 
amino-acids  present  in  lOU  grams  of  liver,  in  ten  days '  autolysis  there 
had  been  set  free  but  1.85  gm.,  after  30  days  10.1  gm.,  and  after  50 
days  but  29.1  gm.  (Abderhalden  and  Prym.**)  By  determining  the 
freezing  point  and  conductivity  of  autolyzing  mixtures,  valuable 
evidence  can  be  obtained  as  to  the  rate  of  change,  which,  in  some  cases, 
is  much  more  significant  than  the  usual  estimation  of  soluble  and  in- 
soluble nitrogen  (Benson  and  Wells. ^)  Titration  of  the  free  amino- 
acids  by  formaldehyde,  together  with  the  estimation  of  proteose  and 
peptone  nitrogen,  also  furnish  valuable  information,  while  the  Van 
Slyke  method  of  determining  free  amino-acids  is  especially  useful  for 
this  purpose. 

Since  Jacoby's  paper  appeared,  the  field  has  been  invaded  by  many 
workers,  who  have  examined  practically  every  tissue  in  the  bod}',  and 
found  that  all  possess  the  power  of  self-digestion ;  or,  in  other  words, 
proteases  are  present  in  every  cell  in  the  body.'^  The  rate  of  digestion 
is  very  different  in  different  organs,  however,  liver  digesting  rapidly 
while  brain  and  muscle  tissue  digest  much  more  slowly,  and  the  auto- 
lytic  activity  varies  under  different  conditions;  thus,  fever  causes  a 
great  increase  in  the  proteolytic  activity  of  the  muscles/  The  char- 
acter of  the  antiseptic  used  modifies  greatly  the  rate,  salicylic  and 
benzoic  acids  giving  the  most  rapid  autolysis,  while  of  non-acid  anti- 
septics toluene  is  perhaps  the  least  inhibitory.  One  of  the  most  im- 
portant factors  in  accelerating  autolysis  is  the  H-ion  concentration 
developing  in  the  tissues.*^  Acidity  acts,  partly,  at  least,  by  so  modi- 
fying the  substrate  that  the  enzymes  can  attack  it,  and  a  very  small 
excess  of  acid  will  destroy  the  enz^mies ;  Bradley  '-^  estimates  this  de- 
structive acidity  at  about  that  concentration  of  H-ions  which  is  indi- 
cated by  methyl  orange  and  Congo  red,  the  maximum  autolysis  being 
obtained  with  an  acidity  at  about  Ph  =  6.00.  A  reaction  approxi- 
mating that  of  blood  (Ph  =  7.4  — 7.8)  reduces  autolysis  to  a  mini- 
mum. A  latent  period  has  been  observed  before  autolysis  in  vitro 
seems  to  begin,  part  of  which  time, may  be  occupied  in  the  develop- 
ment of  sufficient  acidity  to  permit  of  autolysis,  although  Bradley's  ^'^ 

«  Zeit.  phvsiol.  Chem.,  1007   (53),  320. 

7  Jour.  Biol.  Chem.,  1010   (8),  01. 

Ta  Except,  perhaps,  tlic  red  corpuscles  (Pincussohn  and  Roques,  Biocliem.  Zeit., 
1914    (64),   1). 

8  Aronsohn  and  P.lunientlial,  Zeit.  l<lin.  :Nred..  1008    (65),  1. 
8a  See  Morse,  Jour.  Biol.  Chem..  1016   (24),  163. 

9  Jour.  Biol.  Chem.,  1015   (22),  113;   1916    (25),  201. 
9a  Jour.  Biol.  Chem.,  1916    (25),  363. 


84  ENZYMES 

results  indicate  that  it  can  be  accounted  for  largelj^  by  the  time  re- 
quired for  proteolysis  to  proceed  far  enough  to  be  detected  by  chemi- 
cal means. 

The  intracellular  proteases  are  not  altogether  like  either  pepsin  or 
trypsin,  for  they  split  proteins  to  the  simplest  elements,  whereas  pep- 
sin carries  the  digestion  only  to  the  peptone  stage  (under  ordinary 
conditions),  and  unlike  trypsin  their  action  is  most  marked  in  a 
faintly  acid  medium.  Furthermore,  the  cleavage  products  seem  to 
contain  a  much  larger  proportion  of  the  nitrogen  in  the  form  of 
ammonia  and  its  compounds  than  is  tlie  case  with  trj-ptic  digestion, 
because  of  the  presence  of  deaminizing  enzymes  which  split  the  NHg 
groups  out  of  the  amino-aeids  and  purines.  According  to  Bostock  ^^ 
the  greater  the  acidity  the  less  NH.,  is  formed.  It  is  quite  probable 
that  in  autolysis  several  other  intracellular  enzj^mes  are  in  action, 
some  of  which  may  not  be  present  in  pancreatic  or  gastric  juice ;  for 
example,  in  the  liver  is  an  enzjTne,  arginase,  which  splits  the  urea 
radical  out  of  the  arginine  of  the  proteins  (Kossel  and  Dakiu^^), 
and  the  enzymes  which  disintegrate  purines  are  also  absent  from  the 
digestive  juices.  On  the  whole,  however,  the  products  are  quite  simi- 
lar to  those  obtained  by  tryptic  digestion.  To  give  a  concrete  ex- 
ample, Dakin  ^-  detected  in  the  products  of  autolysis  by  the  kidney 
in  acid  solution,  the  following  substances:  Ammonia,  alanine, 
a-aminovalerianic  acid,  leucine,  a-pyrollidine  carboxylic  acid,  phenyl- 
alanine, tyrosine,  lysine,  histidine,  cystine,  hypoxanthine,  and  indole 
derivatives,  including  probably  tryptophane.^-"  The  cleavage  of  sim- 
ple peptids  by  different  tissues  shows  characteristic  differences,  the 
distribution  of  the  enzyme  which  splits  glycyl-tryptophane  having 
been  most  studied.  During  life  the  cells  retain  this  enzyme,  and 
hence  it  appears  in  the  body  fluids  only  when  the  tissues  are  being 
rapidly  disintegrated  (^Mandelbaum).^-*^ 

During  autolysis  tlie  changes  are  by  no  means  limited  to  the  pro- 
teins. Glj'cogen  is  split  into  glucose  very  early,  and  the  sugar  under- 
goes further  changes.  Fats  are  also  split  by  the  lipase,  fatty  acids 
being  found  in  autolyzed  organs.  Reducing  substances  appear,  and 
as  before  mentioned,  numerous  volatile  fatty  acids  are  said  to  be 
produced.  l\Iuch  doubt  exists  concerning  tlie  supposed  formation  of 
volatile  fatty  acids  and  ga.ses  during  autolysis  since  it  was  shown  by 
AVolbach,  Saiki  and  Jackson  ^-^  that  anaerolnc  bacteria  are  almost  in- 
variably present  in  aseptically  removed  dog  livei*s,  for  control  of  auto- 
lysis by  anaerobic  cultures  has  seldom  been   carried  out.     However, 

loBiochem.  Jour.,  1912  (G),  388. 
11  Zoit.  i)liysiol.  Cliom.,  1004    (42).  181. 
12. Jour.  of'pIiysiolof.'A',  100.3   (30),  84. 

i^'-'llio    results    of    autolysis    by    dilTcrent    tissues    are    quite    dissimilar.     See 
Kashiwaliara.  Zcit.  phvsiol.  Clipm.',  1913    (85),  161. 
izcMiincli.  nip(l.  Woeh.,  1914   (61).  461. 
"a  Jour.  Med.  Res.,  1909   (21),  267. 


AUTOLYSIS  85 

there  is  mueli  evidence  that  lactic  acid  is  formed,  and  perhaps  par- 
tially destroyed,  in  autolysis  (Tiirkel,^^  Ssobolew").  Carefully  con- 
trolled experiments  by  Lindemann  ^*^  seem  to  show  that  even  in  the 
absence  of  bacteria,  autolyziiig-  liver  and  heart  can  produce  volatile 
acids,  COo  and  hydrogen.  The  increase  in  fat  described  by  some 
authors  is  probably  only  apparent,  and  due  rather  to  the  liberation  of 
the  fat  from  its  combination  with  the  proteins  so  that  it  is  free  and 
not  "masked,"  as  in  normal  organs. ^^  Lecithin  is  also  decomposed, 
yielding  choline,  but  cholesterol  remains  unchanged  except  for  some 
hydrolysis  of  cholesterol  esters.^^  Creatine  is  changed  to  creatinine 
in  autolyzing  muscle,  and  apparently  both  are  formed  in  autolysis  of 
blood  and  liver. ^^'"^ 

The  nucleo-proteins  seem  to  be  attacked  by  the  autolytic  enzymes, 
as  the  purine  bases  are  prominent  among  the  products  of  autolysis, 
and  in  quite  different  proportions  from  those  obtaining  in  digestion 
of  the  same  tissues  by  other  means.  Apparently  autolytic  enzymes, 
like  trypsin,  attack  the  protein  group  of  the  nucleoproteins,  liberating 
the  nucleic  acids.  These  in  turn  are  attacked  by  specific  enzymes, 
nucleases, ^^  which  liberate  the  purine  bases,  which  are  further  de- 
composed by  specific  enzymes,  guanase,  adenase,  etc.  (See  Chap. 
xxi.) 

It  is  improbable  that  the  intracellular  enzymes  are  merely  pancreatic 
?nzymes  taken  out  of  the  blood  by  the  cells,  because  of  the  differences 
previously  cited ;  furthermore,  Matthes  ^^  found  that  the  liver  retained 
its  autolj^tic  power  after  the  pancreas  had  been  extirpated  (in  dogs), 
and  that  the  autolj^tic  degeneration  of  cut  peripheral  nerves  went  on 
just  the  same,  indicating  that  the  autolytic  enzymes  do  not  owe  their 
origin  to  the  pancreas. 

AVlienever  tissues  are  disintegrated  in  any  considerable  quantities, 
as  after  extensive  burns,  peptolj'tic  enzymes  become  demonstrable  in 
the  blood  and  urine,  and  presumably  these  are  related  to  the  cell  auto- 
lysis.^^*^  They  are  noticeably  increased  in  most  infectious  diseases  in 
which  the  reaction  between  the  body  defenses  and  the  infecting  or- 
ganism takes  i)laee  in  tlie  blood  stream  (Falls). ^°''  Also,  in  the  pre- 
mortal state  a  similar  increase  in  peptolytic  enzyme  in  the  serum  is 

isBioohcm.  Zeit.,  inOO   (20),  431. 

14  Ibid.,  1912  (47),  367.  See  also  v.  Stein  and  Salkowski,  Biochem.  Zeit.,  1913 
(40),   486. 

10  Zeit.  f.  Biol.,  1910   (.55),  36. 

15  See  Krontowski  and  Poleff,  Beitr.  Path.  Anat.,  1914   (58).  407. 

1"  Corper,  Jour.  Biol.  Chem.,  1912  (11),  37;  Kondo,  Biochem.  Zeit.,  1910  (27), 
427. 

17a  Myers  and  Fine,  Jour.  Biol.  Chem.,  1915  (21),  583;  Hoagland  and  McBryde, 
Jour.  Agric.  Res.,  1916   (6),  535. 

IS  Sachs.  Zeit.  physiol.  Chem.,  1905  (46),  337;  Jones,  ibid.,  1903  (41),  101, 
and  1906   (48),  110.* 

19  Arch.  f.  exp.  Path.  u.  Pharm.,  1904   (51),  442. 

loaSee  Pfeiffer,  Miinch  med.  Woch.,  1914   (61),  1099,  1329. 

19b  Jour.  Infect.  Dis.,  1915    (16),  466. 


86  A\zv.i//;,s' 

associated  witli  a  liii:li  iioii-pi'otciii  iiilro^cu  ligure  for  the  serum.^®° 
The  relation  of  the  autolytic  enzymes  to  the  increased  proteolytic 
l)()wer  of  serum,  as  evidenced  in  the  Abderhalden  reaction  {q.v.)  has 
not  yet  been  determined,^'"'  but  Falls  finds  evidence  of  their  correla- 
tion.''"' Blood  proteases  are  also  increased  in  pregnancy.  They 
bear  no  constant  relation  to  the  leucocyte  count. 

Influence  of  Chemicals  on  Autolysis. — As  a  general  rule  the  addition  of  anti- 
seplics  lo  tissues  to  prevent  liaderiai  action  reduces  tlie  rate  of  autolysi.s,  but 
as  most  of  tlie  results  of  "aseptic"  autolysis  so  far  reported  are  open  to  question, 
there  is  a  reasonable  doubt  a.s  to  just  how  much  depression  of  autolysis  there  is. 
Yoshinioto^o  finds  that  of  the  antiseptics  ordinarily  used,  salicylic  acid,  boric 
ai-id,  and  mustard  oil  (one-eiirhth  saturated  solution)  permit  tlie  greatest  auto- 
lysis: but  it  is  probable  that  the  acidity  of  the  lirst  two  antiseptics  plays  an 
ini|)()rtant  ]>art  by  trausforming  pro/ynics  into  en/.ymes  and  by  destroying  in- 
hibiting substances,  hence  the  value  of  the  results  obtained  in  autolysis  with  these 
acids  is  questionable.  However,  sodium  salicylate  and  benzoate  are  said  to 
favor  autolysis  (Laqueur^i).  Toluene  seems  to  interfere  much  less  with  auto- 
lysis than  chloroform  or  thymol  (Benson  and  Wells -2),  and  bromides  are  less 
harmful  tiian  toluene  (Laqueur).  Toluene  vapor,  acting  on  solid  aseptic  tis- 
sues, seems  to  cause  more  depression  of  autolysis  than  is  usually  observed  in 
autolysis  in  solution. 23  Dorothy  Court  -*  found  the  only  satisfactory  antiseptics 
to  be  chloroform,  formaldehyde,  benzoic  and  salicylic  acids,  and  IIXC;  she  em- 
phasizes the  fact  that  for  dillVrcnt  sorts  of  niat(>rials  tiie  diti'erent  antiseptics 
give  variable  results,  so  that  the  antiseptic  used  should  be  selected  with  reference 
to  the  material.  Autolysis  proceeds  rapidly  in  weak  ethyl  alcohol,  5  per  cent, 
being  the  minimum  strength  that  will  prevent  putrefaction:  for  complete  sup- 
pression t)f  autolysis  by  alcohol  the  strength  must  be  at  least  00  per  cent,  net,  after 
allowing  for  the  water  content  of  the  tissues   (Wells  and  Caldwell)  .2-i;i 

Certain  inorganic  substances  in  proper  concentrations  may  increase  the  rate 
of  autolysis  [mercury  2s  and  silver, 20  (colloidal  or  salts)],  yellow  phosphorus, 2" 
iodides, 2s  arsenic, 2!>  CaClo,30  salts  of  Fe,  Mg,  and  cobalt,''i  as  well  as  salts  of 
selenium,  tellurium, 32  and  manganese, 32a  colloidal  sulfur  "^i)  but  not  colloidal 
carbon. 3'.;i-  The  favorable  concentrations  of  these  metals  are  very  low:  thus 
the  o])tim\un  proportion  of  arsenic  is  0.007  milligrams  |)er  1  gm.  tissue,  while  0.04 
mg.  inhibits  autolysis.     ('()._.  increases  and  oxygen  decreases  autolysis  3-ia   jn  litro 

is'cSee  Schul/.  Miiii.l:,  nied.  Wodi.  ]!)l:i  ((iO),  2.)12;  Mandelbaum,  ibid..  1014 
(61),  401. 

ii'«l  See  Sloan,  Anier.  .b>ur.   I'liysiol.,   litl.-)    (30),  0. 
2oZeit.  physiol.  Chem.,  1908   (58),  341. 

21  Zeit.  phVsiol.  Chem.,   1012    (70),  38  and  tif). 

22  .Jour.  Hiol.  Chem.,   1010    (8),  01. 

23  Cruick.shank,  Jour.  Path,  and  Bact.,  1011    (l(i),  107. 

24  Proc.   Rov.  Soc,  Edinburgh,   1012    (32),  251. 
2-Ja,Tour.  Biol.   Chem..   1014    (10).  57. 

2-.  Trufli,  Biochem.  Zeit.,   1010    (23),  270. 

2«Izar,  ihiiL,  1000    (20).  240. 

2i  Sa\l,   llofmeister's  Beitr..   1007    (10),  447;   Vin-liow's  Arch..   1010    i202),   140. 

2*' Kepiiiow.  Hiocheni.  Zeit.,  1011  (37*.  23S :  Kaschiwali;ii-a,  Zeit.  ]ili\si<)l.  Cliein., 
1012    (H2),  -42.'). 

2»  Izar,  Miochem.  Zeit.,  1000  (21),  4();  Laqueur  and  I'.ttiuLier.  Zeit.  phxsiol 
Chem.,   1012    (70).   1. 

aoBriill,  Biochem.  Zeit.,  1010   (20),  408. 

31  Pn-ti.  Zeit.  phvsiol.  Clieiii.,  IMOil  (OOi,  317:  I'niliiii.  I!ineheiii.  Zeit..  1012 
(47).  30(i. 

32  Kasiani,  .\rcli.  sci.  med.,   1012    (30),  430. 

•■"2.1  I'.iadlev,  .lour.   I5i(.l.  (hem..   1015    (21),  200:    1015    (221,   11.3. 
;>2i>  Kagiiioli.    JtiiM-heMi.  Zeit..    101.3    (50).  20  1. 
•«2<-  i/ar  and    I'atane.  ihiil..  \>.  .307. 

•''•''II  .M.  .Moise  found  iiwgen  uithnut  eMeet  mi  aMtul\si>.  Hiocheni.  Uullef  I0l5 
(5).    143. 


RELATION  OF  AUTOrAHfS  TO  M F.TMiOIJSM  87 

(I.aqm'ur).  Thoro  is  disagrooiiient  as  to  vvlietlier  radium  rays  auf,'ment  autolysis. •■'•■* 
Injection  of  iodids  into  animals  is  said  to  increase  the  jjostinortem  autolysis  of 
their  tissues  (Stookey,  Kepinow),  as  also  do  iron  salts,3:ib  while  lar>re  doses  of 
salicylates  decrease  it  (Laciucur).  ]Morse ''■*f  attributes  the  acceleratinj;  action 
of  iodin  and  Itromin  to  increased  acidity  from  formation  of  halogen  acids,  and 
Bradley  "  finds  evidence  that  most  inorganic  salts  that  stimulate  autolysis  act 
by  increasing  Il-ion  concentration.  Addition  of  tuberculin  to  tissues  at  first 
delays  and  tiien  increases  the  autolysis  ( Pe.sci  ;^» ) .  and  dii)htheria  toxin  in  snuiU 
amounts  increases  autolysis  ( Barlocco,-*''  Bertolini  3<i ) ,  neutralization  by  anti- 
toxin not  preventing  this  effect.  Lipoids  also  accelerate  autolysis  (Satta  and 
Fasiani37).  According  to  Soula  ^'a  narcotic  poisons  decrease,  and  convulsive 
poisons  increase  the  rate  of  autolysis  of  nervous  tissue.  Glucose  in  one  per  cent, 
concentration  decreases  autolysis,  and  this  may  be  related  to  the  "protein-sparing 
action  of  carboliydrates."  3Tb  Extracts  of  various  ductless  glands,  or  removal  of 
tliese  glands  from  animals,  seem  to  liave  but  slight  effect  on  autolysis. •'!■<■ 


RELATION  OF  AUTOLYSIS  TO  METABOLISM 
It  liavino-  been  shown  that  proteases  are  present  in  all  cells,  the  next 
question  to  be  considered  is,  do  they  act  only  to  destroy  tissues  after 
death,  or  are  they  of  importance  in  metabolism?  Since  it  is  pre- 
sumably necessary  for  proteins  to  be  split  into  diffusible  and  easily 
oxidized  forms  in  order  that  they  may  enter  the  cell,  and  be  built  up 
into  the  cell  proteins,  or  be  decomposed  with  the  liberation  of  energy, 
the  autolj^tic  proteases  may  be  assumed  to  be  of  prime  importance  in 
protein  metabolism ;  but  to  prove  it  is  another  matter.  Jacoby  found 
that  if  he  lig-ated  off  a  portion  of  the  liver  and  let  it  remain  in  situ 
in  the  animal  the  necrotic  tissues  showed  an  accumulation  of  leucine, 
tyrosine,  and  other  splitting  products  of  the  proteins,  which  suggested 
that  these  same  bodies  are  being  formed  in  the  liver  constantly,  but 
that  they  are  as  constantly  removed  from  the  normal  organs  by  the 
circulating  blood,  or  are  undergoing  further  alterations  which  cease 
when  the  circulation  is  checked.  The  influence  of  various  chemicals 
upon  nitrogen  elimination  seems  to  correspond  to  their  effect  on 
autolysis  (Izar,^^  Laqueitr^^).  Also,  the  histological  changes  of  star- 
vation are  similar  in  many  respects  to  those  of  autolysis  (Casa- 
Bianchi"*°).  Among  other  observations  possibly  bearing  on  the  same 
question  are  those  of  Hildebrandt,"  who  found  that  autolysis  in  the 

33  See  Loewenthal  and  Edelstein,  Biochem.  Zeit.,  1008    (14),  48o :   Brown     \rch 
Int.  Med..  1912    (10),  405. 

33bKottmann.  Zeit.  exp.  Path.,  1912    (11),  355. 
330  .Tour.  Biol.  Chem.,   1915    (22),   125. 

34  Cent.  f.  Bakt.,  1911    (59),  71   and  186. 

35  Cent.  f.  Bakt.,  1911    (60),  43. 

36  Biochem.  Zeit.,  191.3    (48),  448. 

37  Berl.  klin.  Woch..  1910    (47),  1500. 

37a  Conipt.  Bend.  Soc.  Biol.,  1913    (73).  297. 
37b  Shaffer.  Proc.  Soc.  Biol.  Chem.,  1915    (81.  xlii.  . 
37c  Izar  and  Faciuoli,  Sperimentale.  1916    (70).  265. 
3"  Internat.  Beitr.  Erniihrmigstor.,  1910  (1),  287. 
30  Zeit.  physiol.  Chem..  1912    (79).  1  pt  seq. 

40  Frankfurter  Zeit.  Pathol..   1909    (3),  723. 

41  Hofmeister's  Beitr-ige,  1904   (5),  463:  see  also  Grimmer,  Biochem.  Zeit.,  1913 
(53),  429. 


88  ENZYMES 

fuuctionating  inammary  gland  is  much  more  active  than  in  the  rest- 
ing ghmd ;  and  of  Schlesinger/-  wlio  found  tliat  autolysis  was  at  its 
maximum  (in  rabbits)  in  new-born  animals,  decreasing  rapidly  in 
the  fii-st  few  months  of  life,  and  that  in  conditions  associated  with 
emaciation  the  rate  of  autolysis  varied  directly  with  the  degree  of 
enuiciation.  Wells  ■*^  sought  for  a  possible  influence  on  autolysis  by 
thyroid  extract,  which  increases  protein  metabolism,  but  could  demon- 
strate none  in  vitro;  Schryver,*^  however,  found  that  autolysis  was 
more  rapid  in  the  liver  of  dogs  fed  thyroid  extract  for  some  days 
before  death  than  it  was  in  control  animals.  The  results  of  the  former 
observer,  but  not  those  of  the  latter,  have  l)een  confirmed  by  ^lorse.*^' 

DEFENSE  OF  THE  CELLS  AGAINST  THEIR  AUTOLYTIC  ENZYMES 
The  question  of  wliy  the  autolytic  ferments  do  not  destroy  the  cells 
until  after  death  is  a  revival  of  the  old  ]H-oblein  of  "why  the  stomach 
does  not  digest  itself,"  and  the  answer  tliat  satisfies  some  is  that  dead 
protoplasm  is  essentially  different  from  living  protoplasm.  ]More  ex- 
act replies  are  suggested  by  AViener's  studies  on  the  relation  of  the 
reaction  of  the  tissues  to  their  autolysis.  He  found  that  autolysis 
does  not  begin  in  an  organ  until  the  original  alkalinity  is  neutralized 
by  the  acids  which  are  formed  in  all  dead  and  dying  cells.*^  If 
enough  alkali  is  added  to  the  material  from  time  to  time  to  neutralize 
the  acidity  as  it  develops,  autolysis  does  not  take  place.  Although 
the  spleen  contains  an  enzyme  digesting  in  alkaline  solution,  and  an- 
other which  acts  best  in  weak  acids,  the  latter  appears  more  promi- 
nently under  ordinary  conditions  because  the  spleen  and  the  blood 
contain  antibodies  which  check  the  enzyme  that  acts  in  alkaline  solu- 
tions, while  acids  destroy  this  antibody  CHedin).'*^  Organic  acids  are 
formed  in  autolysis  of  the  tissues,  aiul  the  latent  period  between  the 
time  of  the  removal  of  an  organ  from  llic  body  and  the  appearance  of 
autolysis  may  be  explained  partly  by  the  time  required  for  the  neu- 
ti-alization  of  alkaleseeuce.  P)radley  "  has  also  ()l)tained  evidence  that 
the  acid  renders  the  substrate  susceptible  to  digestion  by  the  proteases. 
The  old  observation  that  rigor  mortis  disappears  most  rapidly  in 
nuiscles  that  have  been  exhausted  just  before  death  is  also  probably 
exi)lained  by  the  greater  amount  of  acid  in  such  muscles.  If  we 
imagine  that  autolysis  is  limited  to  periods  when  the  cells  have  an  acid 
reaction,  however,  wo  limit  tlic  i-jiiigc  of  usefidness  in  the  living  cell 

<2  TIofiiK'ifitor'R  Bcitr.,  1003    (4),  !-7 

-•3  Amor.  Jour,  of  Plivsiol.,  1004  (11),  351;  ooi  rolidiatcd  Iiv  l\<itt  maim,  Zeit. 
klin.  ^\^Hl.,  lOin   (71),  3(50. 

4<.Toiir.  of  Plivsiol.,  mn.T  (32),  ir>n. 

■••''.Tonr.  Biol,  riicin.,  lOl.'i    (22),  12r). 

<"  Opifi  (loc  cit.)  found,  liovvovor,  that  autolysis  of  loueocytoa  was  more  rai)id 
in  an  alkalino  incditun.  Docho/,  (Proc.  Sop.  Exp.  Biol,  and  ^I'^d..  1010  (7).  07) 
stat-oH  that  llvfT  alno  contains  an  cn/ynip  active  in  an  alkaliiii'  iiu'diiim.  Ii\it  which 
exists  as  an  inactive  zyniofrcn  until  activated  by  acids. 

••7  Festschrift  f.  liainniarston,  Upsala,  190G. 


DEFENSE  OF  CELLS  AGAIXST  THEIR  AUTOLYTIC  EX/AMES        89 

to  a  minimum,  since  during  life  the  tissue  fluids,  and  presumably  the 
cell  contents,  are  preponderatingly  alkaline.  Perhaps  a  better  ex- 
planation of  the  attack  of  the  cells  by  their  own  enzymes  after  death 
is  to  be  sought  in  the  conditions  of  chemical  equilibrium.  During  life 
constant  new  supplies  of  protein  are  being  brought  to  the  cell,  and  at 
the  same  time  the  products  of  proteolysis  are  presumably  being  car- 
ried away  by  the  circulation  or  being  changed  by  oxidative  processes. 
When  circulation  stops,  the  processes  of  splitting  go  on  without  the 
introduction  of  new  supplies  of  material,  and  hence  the  tissues  are 
not  replaced  as  fast  as  they  are  destroj'ed,  and  the  products  of  their 
decomposition  accumulate,  for  lack  of  any  means  of  destroying  or  re- 
moving them.  The  control  of  autolysis  by  maintenance  of  a  low 
H-ion  concentration  is,  however,  undoubtedly  an  important  factor, 
for  Bradley  ^  found  that  a  reaction  equal  to  that  of  blood  almost  com- 
pletely inhibits  autolysis,  while  the  degree  of  increased  H-ion  con- 
centration that  may  develop  in  local  asphyxia,  or  after  death,  produces 
optimum  conditions  for  autolj^sis. 

Still  another  possible  defense  of  the  living  cells  may  be  found  in 
the  existence  of  specific  antienzymes.  Just  as  the  serum  contains  anti- 
trypsin, so  it  seems  to  contain  substances  antagonistic  to  the  autolytic 
enzjones.  Levene  and  Stookey  found  that  tissue  juices  show  a  resist- 
ance to  digestion,  and  Opie  found  that  the  serum  of  inflammatory 
exudates  retarded  the  action  of  the  autolytic  enzymes  that  are  con- 
tained within  the  leucocytes.  Serum  also  inhibits  autolysis  of  the 
tissues,  so  it  is  probable  that  continuance  of  the  circulation  may  pro- 
vide antibodies  to  the  tissues  to  hold  the  intracellular  enzymes  in 
check,  possibly  without  interfering  with  their  action  on  other  pro- 
teins than  those  of  the  cell  structure.*^*  (See  Antienzymes.)  Lack 
of  oxygen  cannot  be  held  solely  responsible,  according  to  the  studies 
of  Morse,*^''  wdio  found  that  autolysis  occurs  in  muscles  with  divided 
nerves  but  intact  blood  supply.  Nevertheless,  reduced  blood  supply 
results  in  increased  H-ion  concentration  which  greatly  facilitates  auto- 
lysis, and  it  cannot  he  denied  that  autolysis  is  observed  chiefly  if  not 
solely  in  asphyxiated  tissues. 

There  can  be  no  question  that  the  supply  of  food-stuff  is  of  essential 
importance  in  determining  autolytic  changes,  for  it  has  been  found 
by  Conradi,-*^  Rettger.^"  and  Effront  ^'*'  that  bacteria  and  yeasts  begin 
to  undergo  autolysis  when  they  are  placed  in  distilled  water  or  salt 
solution,  which  they  do  not  do,  to  an}-  such  extent  at  least,  when  in 

47a  According  to  Gusrgenheimer  (Deut.  Arch.  klin.  Med.,  1013  {112).  248:  Dent, 
med.  Woch.,  1914  (40),  63),  the  serum  in  various  diseases  has  a  characteristic 
stimulating  or  inhibiting  effect  on  in  vitro  tissue  autolysis,  hut  the  conditions  of 
such  experiments  are  so  complex  as  to  make  their  significance  doubtful. 

47b  Amer.  Jour.  Phvsiol..  101.5   (3fi).  147. 

48  Deut.  med.  Woch.,  1003    (20).  26. 

49  , Jour.  Med.  Research.  1004    (13).  70. 

50  Bull.  Soc.  Chim.,  1005    (33),  847. 


90  f;.v/y.i//;.s' 

iiutriciil  iiiciliii.  (  111  tliis  way  it  has  liccii  iniiiid  ])()ssil)le  to  obtain 
the  intracH'llular  toxins  of  sueli  hactt'ria  as  typlioid  and  cholera.) 
Autolysis  is  not  marked  so  lonj;  as  tlif  bacteria  are  supplied  with 
nourishment.  l)ut  wlicii  nutrient  iiialfiial  is  laekinjz:,  autolytic  decom- 
position is  no  l()ii«ier  repaired  aiul  tiie  bacteria  disintegrate.  Pre- 
sumably the  (•lian<res  are  the  same  in  tissue  cells,  and  anemic  necrosis 
may  be  explained  in  this  way.  Tissue  enxymes  are  also  capable  of 
di<restin«i:  bacteria  (Turro 'M. 

Another  direction  in  which  the  kej^  to  the  action  of  these  enzymes 
may  l)e  sought  has  been  indicated  by  Jacoby,-'"-  who  found  that  to  a 
certain  degree  the  autolytic  enzymes  of  each  organ  are  specific  for  that 
organ.  Liver  extract  will  not  sjilit  lung  tissue,  although  it  will  split 
the  proteoses  that  are  formed  in  lung  autolj'sis,  possibly  because  these 
proteoses  are  less  specific  than  tlic  proteins  from  which  they  arise,  or 
perhaps  because  of  the  erepsin  the  extract  contains  (Yernon). 
Leucocytic  proteases,  however,  seem  capable  of  splitting  foreign 
proteins  of  all  sorts.  Richet  ''^  states  that  the  protease  of  liver  tissue 
does  not  attack  either  muscle  tissue  or  liver  tissue  that  has  been 
coagulated.  Anotlici-  liy])()thesis  has  been  advanced  by  Fermi,"'*  who 
suggests  that  the  ])r()to])lasm  of  living  cells  is  not  digested  because  its 
structural  configuration  is  sucli  that  the  enzymes  cannot  unite  with  it, 
an  attractive  but  practically  undemonstrable  idea. 

Lastly,  it  nuist  be  considered  that  at  least  to  some  extent  the  en- 
zymes exist  in  the  cells  in  their  inactive  zymogen  form,  and  per- 
haps are  changed  into  the  active  foi-m  as  needed,  and  iidiibited 
or  changed  back  again  when  their  work  is  temporarily  finished.  A 
rhythmical  change  of  this  nature  might  be  imagined  as  occurring  and 
accounting  for  interaction  by  the  enzymes,  particularly  since  rhythmi- 
cal changes  in  metabolism  are  known  to  occur  (e.  g.,)  rhythmical  pro- 
duction of  carbon  dioxide  ( Lyon '''•'^ ) .  , 

AUTOLYSIS  IN  PATHOLOGICAL  PROCESSES 

All  absorption  of  dead  oi-  injured  tissues,  and  of  organic  foreign 
))odies,  seems  to  be  accom|)lislied  by  means  of  digestion  by  the  enzymes 
of  the  cells  and  tissue  fluids.  AVe  nmy  distinguish  between  the  diges- 
tion l)rought  about  by  the  enzynu's  of  the  digested  tissue  itself,  or 
autolysis,  and  digestion  by  enzymes  from  other  cells  or  tissue  fluids, 
or  heterohjsis  (Jacoby).  TIeterolysis  is  aeconiplislied  particularly  by 
the  leucocytes,  which  contain  ferments  cajiable  of  digesting  not  only 
leucocytic  proteins  but   a])parently   every  other  sort,''**   from   serum- 

'■'  (inf.   f.   Bakt..   inn2    (.T2),   10."). 
■'•-  IlofiiK'isfoi'K  Bcitr.,   ino.T    (.3).  440. 
f-iC'oini.l.    Itcnd.  Sue.    lUdl..    inn.1    (f).")).   (!.")«. 
f""  (Viil.  f.  Hiikt..  1010   (riC).  .->.^). 
r'-'  Scionro.  1004    MO).  .'{.^O. 

'•"■Many  nulliors  snirp'st  flint  flic  jciicocvf cs  iiicrclv  ciinv  oh/mik's  from  ono 
orpin,    parliciilarly    tlic    pancreas.    f<i    anoflicr.    aiul    lliat    tlicsc    cii/vnics    arc    not 


AUTO/ASIS  J\   l>  \Tlli)l.<)(;i(\L   I'mx'EfSFfEH  91 

iilbuinin  to  catgut  ligatures.  'I'lic  lictci-olysis  iiuiy  be  iutracellular 
■\vliou  the  material  to  be  digested  luis  tirst  beeu  taken  up  by  the  cells 

(phagocytosis)  ;  or  extra-cellular,  either  by  enzymes  normally  con- 
tained in  the  blood  ])lasma  and  tissue  fluids,  or  by  enzymes  liberated 
by  the  leucocytes  and  tixed  tissue  cells.  On  death  and  dissolution  of 
a  cell  the  intracellular  enzj'mes  are  released,'*"^  but  it  is  not  known  to 
what  extent  the  enzymes  may  be  secreted  from  intact  living  cells.  As 
far  as  pathological  processes  show,  the  amount  of  liberation  of  en- 
zymes from  normal  cells  is  very  slight,  if  any,  and  the  digestive  en- 
zymes of  the  blood  plasma  seem  to  be  very  feeble,  but  this  is  perhaps 
because  they  are  largely  held  in  check  by  the  anti-enzymatic  substances 
of  the  serum.  I'athological  autolysis  and  heterolysis,  therefore,  are 
brought  about  chiefly  by  enzymes  liberated  from  dead  or  injured  cells. 
Bacteria,  however,  can  multiply  upon  a  medium  of  coagulated  protein, 
M'hich  suggests  that  they  also  secrete  proteolytic  substances,  for  other- 
wise it  would  be  difficult  to  explain  how  they  secure  their  nourish- 
ment. In  pathological  conditions,  digestion  of  degenerated  tissues 
seems  usually  to  be  the  result  of  both  autolysis  and  heterolysis.  An 
infarct  softens  because  the  intracellular  enzymes  digest  the  dead  cells, 
exactly  as  they  do  when  the  tissue  is  removed  from  the  body,  ground 
up,  and  put  in  the  incubator  under  toluene.  In  addition  leucocytes 
wander  in,  disintegrate,  and  their  liberated  enzymes  help  in  the  proc- 
ess, as  also  do,  to  a  less  degree,  the  enzymes  of  the  blood  plasma.  It  is 
because  of  the  heterolysis  by  leucocytic  enzymes  that  a  septic  infarct 
becomes  softened  so  much  more  rapidly  than  does  a  sterile  infarct,  and 
by  comparing  the  rate  of  softening  in  septic  and  aseptic  infarcts  we 
see  that  the  cellular  autolysis  is  a  very  slow  process  as  compared  to 
the  heterolysis  accomplished  by  the  leucocytes.  The  explanation  of 
this  may  lie  in  the  fact  that  most  intracellular  proteases  act  best  in 
an  acid  medium  (Wiener),  while  leucocytic  proteases  act  best  in  an 
alkaline  medium  (Opie),  and  the  infarcts  of  small  size  are  seeped 
through  by  alkaline  blood  fluids.  When  an  infarct  is  large,  we  find 
it  undergoing  central  softening  while  the  periphery  remains  firm ;  this 
corroborates  our  hypothesis,  for  acids  are  developed  during  autolysis 

(Magnus-Levy),  which  at  the  periphery  are  neutralized  by  the  blood 
plasma,  so  that  only  at  the  center  is  autolysis  active.     The  inhibiting 

formed  by  the  leuooevte  itself.  Opie  (Jour.  Exp.  ^led.,  10(1.7  (7).  T.IO )  Juis  shown, 
liowever.  that  the  bone-marrow  contains  proteolytic  enzymes  which  are  like  those 
of  the  leucocytes  in  that  tliey  act  best  in  an  alkaline  medium,  whereas  the 
autolytic  enzymes  of  the  lymphatic  tflands  and  most  otliei-  tissues  act  best  in 
an  acid  medium.  This  leaves  little  room  for  doubt  that  the  leucocytes  are 
equipped  with  their  characteristic  enzymes  when  they  leave  the  bone-marrow, 
and  that  they  are  not  obtained  later  in  the  pancreas  or  elsewhere.  ^Fore  re- 
cently, however,  van  Calcar  (Pfliijier's  Archiv.,  1012  (148),  2rt~)  has  revived  the 
idea  of  the  origin  of  leucocytic  enzymes  in  the  digestive  ^rlands. 

ooa  Peptolytic  enzymes  appear  in  the  urine  after  severe  superficial  burnincr,  pre- 
sumably comins:  from  the  disintegrated  cells.  (Pfeiffer,  INIiinch.  med.  Wocli  , 
1914   (61),  1329.) 


92  ENZYMES 

action  of  the  serum  also  has  a  similar  effect,  limiting  autolysis  at  the 
periphery. 

In  the  case  of  septic  softeniny;  the  action  of  the  bacteria  needs  also 
to  be  taken  into  consideration,  since  they  also  produce  proteolytic 
ferments,  but  their  effect  seems  to  be  relatively  small  as  compared  with 
leucocytic  digestion.  Intracellular  digestion  of  necrotic  tissue  by 
leucocytes  seems  also  to  be  relativelj^  unimportant.  Suppuration, 
therefore,  must  be  considered  as  the  result  of  digestion  of  dead  tissue 
by  enzymes  derived  from  the  leucocytes,  the  plasma,  the  bacteria,  and 
the  destroyed  cells  themselves.  A  tubercle  does  not  ordinarily  sup- 
purate, because  the  tul)er('le  bacillus  and  the  substances  it  produces 
are  not  strongly  chemotactic,  and  hence  not  enough  leucocytes  enter 
the  necrotic  area  to  produce  a  digestive  softening.  The  enzymes  of 
stai)liylo('Occus  are  much  more  strongly  proteolytic  than  those  of 
streptococcus  (Knapp^'),  which  may  be  one  reason  why  the  latter  so 
much  more  frequently  produces  lesions  without  suppuration  than 
does  the  former.  Necrotic  areas  of  any  kind  are  absorbed  by  similar 
processes.  Autolysis  of  tumors  is  quite  active  in  specimens  removed 
from  the  body,  and  the  areas  of  necrosis  that  occur  commonly  in 
tumors  are  absorbed  in  this  way.  Apparently  all  varieties  of  cells 
are  subject  to  autolj'sis  or  heterolj^sis  whenever  they  are  killed  or 
sufficiently  injured.  Involution  of  the  uterus  probably  depends  upon 
autolj'sis,  which  is  much  more  active  in  the  puerperal  uteinis 
(Ferroni^^),  and  creatine  is  found  in  the  urine  when  such  autolysis 
occurs,^^  although  A.  INIorse  -'^  considers  this  to  be  independent  of 
the  uterine  autolysis.  Atrophy  may  be  looked  upon  as  an  autolysis 
in  the  normal  course  of  catabolism,  not  met  by  a  corresponding  build- 
ing up  of  the  proteins,  l)ut  ]\I.  ^Morse  ^''''^  could  tind  no  evidence  that 
the  atrophy  and  involution  of  the  tadpole  tail  is  accompanied  by  an 
accelerated  autolysis.  The  solution  of  fibrin  by  tissues,  fihrinolysis, 
is  considered  to  be  distinct  from  tissue  autolysis  by  Fleisher  and 
Loeb.'"''"'  In  atrophic  cirrhosis  the  fibrinolytic  activity  of  the  blood  is 
increased,  wliidi  may  explain  the  lieniorrliagic  tendency  of  this  dis- 
ease.^®° 

The  products  of  autolysis  may  of  themselves  be  toxic;  albumoses 
and  pe])tones  certainly  are,  and  the  other  cleavage  products  are  prob- 
abh^  not  altogether  innocuous.  (See  "  Autoinloxication.")  Some  of 
the  symptoms  of  suppuration,  ])articularly  llic  fever  and  chills,  have 
been  ascribed  to  the  autolytic  products  i-atlni-  tlian  to  the  bacterial 

"Zcit.  f.  ITeilk.    (riiir.).  1902    (2.3).  2.30. 

f'SAnn.  (li  Ostctrica  e.  Ginocol.,  IflOfi  (2),  .'i5.3:  set'  also  Slciuons,  Bull.  Johns 
Hopkins  Ifosp.,  1014  (2.'5),  195;  Arthur  Morso,  Jour.  Amer.  Med.,  Assoc,  1915 
(0-)),   1(11.3. 

so  SliiifTor.  Amor.  .lour.  I'hvsiol.,   190S    (23).   1. 

snnMiix  Morso,  Am.  .lour.  I'hvsiol.,   lOl.'i    (3G),  14r). 

si'b.Tour.  IJiol.  Chom.,   IfM.'}    (21).  477. 

'•!'■•  Coodpnslurc,  I'.uli.  .lolins  Hopkins  ITosp.,  1914   (25),  330. 


AUTOLYSIS  IX  I'ATUOLOGICAL  PROCESSES  93 

poisons,  particularly  as  aseptic  suppuration  is  accompanied  by  fever, 
Joclimanu  '^'^  lias  found  evidence  that  the  protease  of  leucoytes  can 
cause  fever  and  also  reduce  the  coagulability  of  the  blood.  The  work 
of  Vaughan  and  other  recent  students  of  the  reaction  to  foreign  pro- 
teins, shows  that  typical  fevers  can  be  produced  by  the  enzymatic  dis- 
integration of  proteins  in  the  body."'^''  Degenerative  changes  in  nerv- 
ous tissue  are  associated  with  autolytic  decomposition  of  the  lecithin 
(NoU"^)  and  the  liberated  choline,  or  its  more  toxic  derivatives,  may 
be  a  source  of  intoxication.''-  In  all  conditions  associated  with  auto- 
lysis, such  as  resolving  pneumonic  exudates,  large  abscesses,  softening 
tumors,  etc.,  albumoses  (and  peptones?)  may  appear  in  the  urine. 
Autolytic  products  may  also  be  hemolytic  (Levaditi  "^),  and  thej^  may 
prevent  clotting  of  the  blood  (Conradi  "*).  It  is  probable  that  among 
tlie  products  of  autolysis  are  bactericidal  substances,"^'^  although  it  is 
doubtful  if  the  concentration  is  often  sufficient  for  them  to  be  of 
influence  except  in  well  walled  areas. 

Work  has  been  reported  upon  autolytic  processes  in  a  number  of 
pathological  conditions,  which  may  be  discussed  briefly  as  follows: 

Exudates. — The  presence  of  leucine,  tyrosine,  proteoses,  and  pep- 
tones in  pus  has  been  known  for  many  years,  and  the  reason  for  their 
appearance  is  now  clear.  Miiller,^^  many  years  ago,  observed  that 
purulent  sputum  digested  fibrin,  but  that  non-purulent  sputum  did 
not  have  this  property.  Achalme  "*'  found  that  pus  would  dissolve 
gelatin,  fibrin,  and  egg-albumen.  Ascoli  and  jMareschi  ®"  detected 
autol,ysis  in  sterile  exudates  obtained  experimentally.  Umber  ^^ 
found  that  ascitic  fluid  exhibited  autolytic  changes,  which  observa- 
tion could  not  be  confirmed  by  Schiitz  ^°  in  pleural  exudates  and  as- 
citic fluids.  Zak  ^°  found  that  autolysis  was  inconstant  in  various 
exudates.  The  ditferences  in  these  results  are  explained  by  Opie's"^ 
observation  that  in  experimental  inflammatory  exudates  the  leuco- 
cytes are  capable  of  marked  autolysis,  whereas  the  serum  contains  an 
antibody  which  holds  this  autolysis  in  check;  if  the  antibody  is  de- 
stroyed by  heat,  then  tlie  serum  proteins  are  also  digested  by  the  leu- 
eocytic  enzymes.     This  antibody  seems  to  be  contained  normally  in 

GoVirchow's  Arch..   1908    (104).   342. 

60a  See  Vaughan,  "Protein  Split  Products."  Philadelpliia.   101.3. 

Gi  Zeit.  physiol.  Chemie,    1809    (27),  380. 

G2  See  Haliibvirton,  Erpebnisse  der  Plivsiol.,  1904   (4),  24. 

G3  Ann.  d.  I'Inst.  Pasteur,  1903  (17),  187;  also  Fukuhara.  Zeit.  f.  exp.  Path.  u. 
Pharm..  1907    (4),  658. 

64  Hofmeister's  Beitr.,   1901    (1),   130. 

64a  See  Bilancioni,  Arch,  farmacol.,   1911    (11),  491. 

GsKossel,  Zeit.  f.  klin.  ]\Ied..  1S8S    (13),  149. 

««  Compt.  Rend.   Soc.   Biol.,   1899    (.51),  568 

G7  See  Malv's  Jahresbericht.   1902    (32),  568. 

osMiinch.  ined.  Woch.,  1902    (49).  1169. 

69  Cent.  f.  inn.  Med.,  1902   (23),  1161. 

70Wien.  klin.  Woch.,   1905    (18),  376. 

71  Jour,  of  Exper.  Med.,  1905  (7),  316  and  759;  1906  (8),  410  and  536;  1907 
(9),  207,  391  and  414;  also  a  full  review  in  Arch.  Int.  Med.,  1910   (5),  541. 


94  EXZYMES 

the  albumin  of  the  blood-serum.  In  old  exudates  the  antibodies  are 
decreased,  and  autolysis  then  occurs,  explaining  the  variable  results 
of  Umber,  Schiitz  and  Zak.  The  intracellular  proteases  of  the  poly- 
nuelear  leucocytes  act  best  in  an  alkaline  medium;  those  of  the 
mononuclears  in  an  acid  medium.  If  the  proi)()rti()n  of  serum  to 
leucocytes  is  high,  then  there  is  no  autolysis,  as  in  serous  exudates; 
but  if  the  leucocytes  are  abundant,  then  the  antibody  is  overcome 
and  we  get  autolysis,  as  in  ordinary  suppurative  exudates.  Animals 
with  but  little  protease  in  their  leucocytes  (e.  g.,  rabbits),  do  not 
ordinarily  produce  a  liquid  pus  (Opie).  Exudates  produced  by  bac- 
terial infection  also  seem  to  possess  the  properties  above  described. 
Galdi  '-  found  autolysis  greater  in  exudates  than  in  transudates,  but 
observed  no  constant  relation  between  the  number  of  leucocytes,  or 
the  amount  of  chlorides,  and  the  rate  of  autolysis.  All  exudates,  ac- 
cording to  Lenk  and  Follak,''  contain  enzymes  splitting  glycyl-gly- 
cine  (peptolytic  enzymes)  ;  the  most  active  exudates  are  those  of 
cancer  and  tuberculosis,  the  least  active  are  passive  congestion  fluids; 
pleural  exuchites  contain  more  active  enzymes  tlian  peritoneal  exudates 
of  similar  character. 

Knapp  '*  holds  that  in  pus  the  cocci  and  tlie  enzymes  they  produce 
are  responsible  for  much  of  the  digestion.  Pus  cells  alone  do  not 
undergo  digestion  so  rapidly  as  wlien  bacteria  are  present,  and  di- 
gestion is  more  rapid  if  the  bacteria  are  alive  than  when  inhibited  or . 
killed  by  antiseptics.  Streptococcus  is  almost  inactive,  staphylococcus 
is  quite  active,  and  B.  coli  still  more  so.  However,  pus  corpuscles 
free  from  bacteria  are  highly  proteolytic,  causing  digestion  in  serum 
plates  in  dilutions  of  1-700  (Jochmann).  Knapp  could  find  no  rela- 
tion between  the  autolytic  power  of  the  pus  and  the  severity  of  the  in- 
fection from  which  it  resulted.  A  constant  constituent  of  pus  is 
d-lactic  aeid,^^^  and  it  increases  during  autolysis ;  this  may  well  modify 
the  rate  of  autolysis  of  pus.  (See  also  tlie  discussion  of  the  "Chem- 
istry of  Tiis, "  Cha]).  X.) 

Proteolytic  Enzymes  of  the  Leucocytes.'-' — By  tlie  introduction  of 
the  plate  method  of  testing  the  proteolytic  activity  of  leucocytes, 
Miiller  and  Jochmann  brought  the  study  of  this  particular  vital 
activity  into  the  range  of  clinical  laboratories,  and  aroused  nuu-h 
general  interest  in  what  had  previously  concerned  only  a  few  pathol- 
ogists, especially  E.  L.  Opic  The  jn-inciple  is  that  of  permitting 
the  leucocytes  or  other  cells  to  act  ui)oii  a  blood  .serum  plate  at  a  tem- 
pei'ature  of  55'^',  which  prevents  bacterial  action  but  permits  the  pro- 

7-' See  Folia  llcmat.,   1!)().")    (2),  .")2n. 

73  Dout.  Arcli.  kliii.  Med..  1013  (100),  ViTiO:  see  iilso  Wiener.  Biocliem.  Zeit., 
1012    (41),   140;   Miuidclliainii.  :Miiiieli.  ined.  Woeli.,   l!tl4    (CI).  4(il. 

7*  Zeitsclir.  f.  lleilk.,   I!l(i2    (2:5.  Cliir.  .Mil.),  2:i(i. 

7-ialto,  .Tour.  ]?i<.l.   Ciieiii..   101(1    (2(1).    17.S. 

"■'•  i"'ull  liililio^'riipliv  liv  \\ieTis,  Kr<relmisse  riivsiol..  lOll  (l.">),  1:  ■locliinaiin, 
Koiie  and  Wasserniaiin's   Ilandliueli.   1012    (2).   1:301. 


AUTOLYSIS  I\  I'ATUOLOaiCAL  I'h'OCESSES  95 

toolytic  on/ymes  of  the  cells  to  (Ii<rt'st  the  coajiulated  serum,  forming 
depressions  in  the  surface  ("  Dellbilduiig").  This  proteolytic  activ- 
ity is,  of  course,  heterolysis  rather  than  autolysis.  ^lany  modifica- 
tions of  this  method  have  been  introduced  (such  a-s  using  casein- 
agar),  but  the  i)rincii)le  involved  is  the  same,  and  they  are  fully  ex- 
plained and  discussed  in  the  article  by  AViens.  Normal  blood  does 
not  contain  enough  leucocytes  to  cause  observable  digestion,  but  my- 
elogenous leukemia  blood  causes  distinct  digestion  while  lymphatic 
leukemia  does  not,  showing  that  it  is  the  polynuclears  and  myelocytes 
that  are  responsible.  Other  observations  fasten  the  proteolytic  activ- 
ity upon  the  neutrophile  granules.  Leucocytes  of  normal  human 
blood  will,  if  concentrated  enough,  cause  digestion  of  serum  plates, 
as  also,  of  course,  will  pus.  The  leucocytes  of  rabbits,  guinea  pigs, 
and  practically  all  animals  except  man,  apes  and  monkeys,  are  de- 
void of  proteolytic  activity  demonstrable  by  the  plate  method.  Nor- 
mal serum,  both  homologous  and  heterologous,  exercises  a  strong  in- 
hibition on  this  digestion,  so  that  it  is  necessary  to  have  an  excess  of 
leucocytes  present  to  obtain  the  reaction.  The  leucocytic  enzymes 
seem  to  be  very  resistant  against  chemicals,  especially  against  formal- 
dehyde, so  that  museum  specimens  of  leukemic  tissues  preserved  in 
formalin  for  years  are  still  proteolytic.  Liver  tissue  is  but  slightly 
proteolytic  by  this  test,  spleen  more  so,  and  leucocyte-containing  flu- 
ids, such  as  saliva  and  colostrum,  are  quite  active.  Pancreas  tissue 
has,  of  course,  strong  proteolytic  action,  but  it  is  shown  to  be  distinct 
from  the  leucocytic  protease  by  being  inhibited  by  certain  sera  that 
do  not  inhibit  the  leucocytic  protease.  In  general,  tissues  do  not 
cause  much  proteolysis  of  serum  plates  unless  they  are  invaded 
by  many  leucocytes,  which  applies  also  to  tumors,  including  mul- 
tiple myelomas.  Besides  proteases,  leucocytes  contain  other  en- 
zj^mes.^® 

To  quote  the  summary  by  ^Morris  and  Boggs,"  "it  has  been  shown 
that  the  normal  and  pathological  neutrophile  leucocytes  and  myelo- 
blasts contain  an  oxidase  and  probably  a  lipase  and  an  amylase; 
myeloblasts  contain  an  amylase.  In  lymphoid  tissues  two  proteases 
and  a  lipase  have  been  shown  to  exist.  In  leukemia  leukoprotease  has 
been  demonstrated  in  the  myeloid  variety  of  the  disease,  while  it  has 
not  been  found  in  chronic  lymphoid  leukemia.  Lipase  has  been  dem- 
onstrated in  two  cases  of  myeloid  leukemia,  and  oxidase  in  all  myeloid 
cases  observed  in  which  the  neutrophilic  cells  were  present  in  excess. ' ' 
Jobling  and  Strouse,'^  confirming  Opie's  observation  of  two  distinct 

"6  Accordino^  to  Tschernoruzki  (Zeit.  physiol.  Cliem.,  1011  (75).  21(i)  amylase, 
diastase,  catalase,  peroxidase,  and  nuclease,  but  not  lipase.  T  also  found  uricase 
absent  from  dog  leucocytes  (Jour.  Biol.  Chem.,  1909  (G),  321).  Fiessin»er  and 
]\Iarie  (Compt.  Rend.  Soc.  Biol.,  1909  (67),  177)  state  that  the  lymphocytes 
contain  lipase,  although  myeloid  cells  do  not. 

77  Arch.  Int.  Med.,  1911*  (S),  806. 

7s  Jour.  Exp.  Med.,  1912   (16),  269. 


96  EXZYMES 

proteases  in  leucocytes,  find  also  evidence  of  an  ereptic  enzyme  act- 
inpr  in  either  Scid  or  alkaline  fluids. 

Pneumonia. — In  the  stage  of  resolution  lobar  pneumonia  presents 
a  striking  example  of  autolysis.  The  often-remarked  phenomenon 
that  the  lung  tissue  itself  is  not  in  the  least  afiPected,  while  the  dense 
contents  of  the  alveoli  are  rapidly  dissolved  and  removed  is  explained 
by  the  invariable  immunity  of  living  cells  to  digestive  enzymes.  Ex- 
cept for  some  slight  possible  assistance  by  the  alveolar  epithelium 
and  the  enzymes  of  the  serum,  the  enormous  and  rapid  digestion  of 
pneumonic  exudates  is  accomplished  by  tlie  leucocytic  enzymes.  The" 
rapid  rate  of  digestion  may  be  accounted  for  bj-  the  absence  of  circu- 
lation within  the  alveolar  contents,  which  permits  the  leucocytes  to 
act  unimpeded  by  the  anti-bodies  of  the  blood  plasma.  Digestion 
of  the  exudate  continues  after  death,  accounting  for  the  marked  dif- 
fuse softening  observed  in  pneumonic  lungs  in  bodies  kept  some  days 
before  autopsy.  As  long  ago  as  1888,  Kossel  ''^  mentioned  that  Fr. 
]\Iiiller  had  found  that  glycerol  extracts  of  purulent  sputum  exhib- 
ited a  digestive  action  upon  fibrin  and  coagulated  protein,  whereas 
non-i)urulent  sputum  did  not  possess  this  property.  In  1877  Filehne 
•extracted  ferments  in  the  same  way  from  the  sputum  in  gangrene 
of  the  lung;  Stolniknow,  in  1878,  found  a  similar  ferment  in  pneu- 
monic sputa,  and  Escherich  in  1885  showed  that  the  proteolytic  action 
of  tuberculous  sputum  was  independent  of  putrefaction.  Other  early 
observations  of  similar  nature  are  reviewed  by  Simon,^°  who  demon- 
strated the  presence  of  leucine  and  tyrosine  in  the  autolyzed  lungs. 
In  a  later  work  IMiiller  reports  finding  three  grams  of  leucine  and 
tyrosine  in  a  pneumonic  lung,  as  well  as  lysine,  histidine,  and  purine 
bases  from  the  decomposed  nucleoproteins.  The  appearance  of  free 
purines  during  autolysis  of  pneumonic  lungs  has  been  investigated 
by  Mayeda,^^  Long  and  Wells.^^''  Boehm  ^-  isolated  histidine  and 
arginine  from  the  same  material.  Rietschel  and  Langstein  ®^  found 
0.32  gm.  leucine  in  the  urine  of  a  pneumonic  child.  Flexner  ^^  noted 
that  autolysis,  while  \ery  rapid  in  the  gray  stage,  is  but  slight  in  the 
red  stage  (because  of  paucity  of  leucocytes)  and  also  in  unresolved 
pneumonia,  which  he  considers  as  due  to  some  interference  with  auto- 
lysis. Silvcstrini  ^^  found  tliat  in  gray  hejiatization  the  reaction  was 
strongly  acid,  in  red  faintly  so;  the  gray  hepatization  showed  more 
peptone,  and  leucine  and  lactic  acid  were  botli  demonstrable.  A 
fibrin-digesting    enzyme    was    isolated,    and    u\\\k    was    coagiilated. 

70Zcit.  f.  klin.  :\rcd.,  18SS    (1.3),   149. 

80  Dent.  Arch.  klin.  Med.,  inOl    (70),  G04. 

81  Dent.  Arcli.  klin.  Med.,  1910   (98),  5cS7. 
8ia/?;tU,  1914   (115),  377. 

»2jhid.,  1910   (98),  .-iS.^. 
83Biodicm.   Zcit.,    1906    (1),   75. 

84  Univ.  of  Penn.  Med.  Bull.,  190.3   (16),  185. 

85  Bull.  del.  Soc.  Kustaeliiana,  1903,  abst.  in  Biochem.  Centralbl..  1903   (1).  713. 


Arroi.Yi^is  J\  rATifOLdcicM.  i'i;(>rEsi<Ei<  97 

Kzentkowski  ^-'  found  an  increase  of  non-i-oayulablc  nitroficu  in  the 
Wood  of  i)n('uni()iii('s,  probably  resulting  from  autolysis  in  the  exu- 
date. Aecordin<r  to  Dick"*'  the  blood  serum  after  the  crisis  contains 
an  enzyme  which  acts  specifically  on  the  pneumocoecus  proteins.  In 
the  liver  during  experimental  pneumocoecus  septicemia,  autolysis  is 
increased  in  rate.''''"'  Almagia  ^'^  suggests  that  the  bactericidal  action 
of  the  jiroducts  of  fibrinolysis  in  ]nieumonia  may  be  of  importance  in 
checkinu'  the  disease. 

Necrotic  Areas. — lacoby  '"'"  found  that  if  a  poi-tion  of  a  dog's  liver 
Avas  ligated  oft*  and  tlie  animal  kept  alive  for  some  time,  the  necrotic 
ti.ssue  contained  tlie  same  jiroducts  that  he  had  obtained  in  experi- 
mental autolysis.  The  absorption  of  necrotic  tissues  generally  is 
ascribable  to  either  autolysis  or  heterolysis.  Presumably  there  is  no 
great  difference  in  the  self-digestion  of  an  organ  which  is  necrotic 
because  its  blood  supply  is  cut  off,  and  of  a  similar  organ  removed 
from  the  body  aseptically  and  allowed  to  undergo  aseptic  autolysis 
in  an  incubator.  At  the  periphery  there  might  be  some  effects  pro- 
duced in  vivo  by  the  inhibitive  action  of  the  serum  or  the  digestive 
action  of  the  leucocytes,  but  beyond  that  no  marked  differences  are 
to  be  expected.  In  both  cases  asphyxia  is  present,  leading  to  increased 
acidity,  without  wliich  little  if  any  autolysis  can  occur. 

A  study  of  the  relation  of  autolysis  to  the  histological  changes  that 
occur  in  necrotic  areas  by  Wells  ^^  gave  evidence  that  there  occurs 
early  a  decomposition  of  the  nucleoproteins  of  the  nuclei,  which  is 
probably  brought  about  by  the  intracellular  autolytic  enzymes.  The 
liberation  of  the  nucleic  acid  and  the  reduction  in  the  bulk  of  nuclear 
material  through  the  digestion  away  of  the  protein  is  probably  the 
cause  of  the  pycnosis  observed  in  necrotic  areas.  Later  the  nucleic 
acids  are  further  decomposed  through  the  special  enzymes  described 
by  Jones,  Sachs,  and  others,  the  "nucleases."  This  is  presumably 
the  cause  of  the  loss  of  nuclear  staining  so  characteristic  of  necrosis. 
That  these  changes  are  due  to  the  intracellular  enzymes  was  shown  by 
implanting  in  animals  pieces  of  sterile  tissues,  the  enzATnes  of  which 
had  been  destroyed  by  heating;  these  were  found  to  undergo  altera- 
tions only  after  several  weeks,  and  then  as  the  result  of  the  action 
tipon  them  of  invading  leucocytes.  The  slow  rate  of  autolj-sis  that 
occurs  in  infarcts  and  other  aseptic  areas  is  presumably  due  in  part  to 
the  action  of  the  antibodies  of  the  serum,  for  it  was  found,  experimen- 
tally, that  the  histological  clianges  of  autolysis  when  the  tissues  are 
placed  in  heated  serum  proceed  about  twice  as  rapidly  as  w^hen  they 
are  placed  in  fresh  serum.     Chemotactic  substances  do  not  seem  to 

SGVirchow's  Arch.,   1005    (179).  405. 
ST  .Jour.  Infect.  Dis..  1912    (10),  .383. 
8"a  ^ledigreceanii.  .Jour.  Exp.  :Med..  1914    (19),  31. 
8"b  Festsclir.  for  Celli,  Torino,  191.3.  p.  459. 
ssZeit.  physiol.  Cliem..   1900    (30),   149. 
89  Jour.  Med.  Research,  1906    (15),   149. 
7 


98  ENZYMES 

be  fonned  in  aseptic  dead  tissues,  but  the  slow  absorption  of  such 
tissues  is,  however,  finally  accomplished  by  the  leucocytes  acting 
from  the  periphery,  there  beino:  little  actual  autolysis  of  the  dead 
cells  by  their  own  enzymes.  Tlie  rapidity  with  which  autolytic 
changes  occur  in  different  organs,  as  indicated  by  the  disappearance 
of  nuclear  staining,  seems  to  be  about  as  follows:  (1)  Liver,  kidney 
^epithelium  of  convoluted  tubules)  ;  (2)  spleen,  ])ancreas;  (3)  kidney 
(collecting  tubules,  straight  tubules,  glomerules)  ;  (4)  lung  (alveolar 
and  bronchial  epithelium)  ;  (5)  thyroid;  (6)  myocardium;  (7)  volun- 
tary muscle;  (8)  skin  (epithelium);  (9)  brain  (cortical  cells). 
Stroma  cells  seem  to  be  attacked  chiefly  by  enzymes  from  the  paren- 
chynui  cells.  Of  all  cellular  elements,  the  endotlielium  of  the  vessels 
seems  to  have  the  greatest  resistance  to  both  autolysis  and  heterolysis. 

The  finer  structural  changes  of  aseptic  autolysis  of  liver  in  salt  so- 
lution, have  been  carefully  studied  by  Launoy,**"  who  notes  a  period 
of  relative  latency  (20  to  2-4  hours  at  38°),  followed  by  rapid  changes 
in  both  cytoplasm  and  nucleus,  associated  with  the  appearance  of 
myelin  forms.  Dj^son ''^  describes  loss  of  the  Altmann's  granules  in 
autolyzing  cells.  Cruickshank  ^-  states  that  when  aseptic  autolysis 
of  tissues  kept  in  a  moist  chamber  is  observed  microscopically  the 
changes  are  slower,  and  there  is  less  solution  of  the  cytoplasm,  but  in 
general  the  results  are  much  the  same.  No  fat  could  be  found  by 
special  stains.  Fetuses  that  have  undergone  aseptic  autolysis  in  the 
uterus  show  complete  loss  of  nuclei  in  5  to  6  days,  a  stage  correspond- 
ing to  8  to  15  days  autolysis  in  the  moist  chamber.  In  experimental 
nephritis  Simons  ^^'^  observed  a  decreased  autdlj-sis  of  the  kidneys. 

Degenerated  nervous  tissue  also  undergoes  a  slow  autolysis  which, 
according  to  Noll,"'*  results  in  the  splitting  of  protagon  with  libera- 
tion of  lecithin.  IMott,  Halliburton.-'''  Donath,  and  others  have  shown 
that  in  nerve  destruction  lecithin  is  split  up  with  liberation  of  cho- 
line (see  "Choline").  Koch  and  Goodson  **"  found  that  degenerated 
nervous  tissue  is  characterized,  chemically,  by  containing  a  relatively 
increased  amount  of  nucleo-proteins,  with  an  absolute  decrease  in 
s'olid  constituents,  while  the  lecithins  a)"e  greatly  altered. 

In  caseation  autolysis  is  very  slight,  as  is  sliown  by  the  persistence 
of  the  caseous  material  for  long  periods  of  time  witliout  absorption. 
Presumably  the  toxin  of  tuberculosis  destroys  the  autolytic  ferments 
of  the  cells  it  kills,"'  and  as  thei-e  is  little  chemotactic  influence,  leuco- 

00  Ann.  Inst.  Pasteur,  1009   (2:^).  1. 

91. Tour.  Patli.  and  l?act.,  HU'J  (17),  12;  also  AsdiolV,  Vcrli.  d.Mit.  Path. 
Gesellscli,   1014    (17),   100. 

92  .lour.  Patli.  and  Pact.,  1011    (10),  1G7. 

92a  Piodiom.  Zoit.,  1014    (67),  483. 

94Zcit.  pliysiol.  Clieni..   1800    (27),  390, 

05  General  r^^'sum^'  in  Kr<,'elinissc  der  Plivaiol.,  1004    (4),  24. 

98Amer.  Jour.  Pliysiol.,  lOOfi    (l.")),  272.' 

07  However,  Pesoi  "(Pa1ii(d()-ica,  1012  CM.  144)  slates  tlial  liilierculiii  increases 
autolysis  in  vitro. 


AUTOLYSIS  IN  PATHOLOGICAL  PROCESSES  99 

cytes  do  not  enter  tlic  caseous  area.  Jobling  and  Petersen  "^'^  find 
evidence  that  the  soaps  of  unsaturated  fatty  acids  present  in  tubercles 
are  responsible  for  the  inliibition  of  digestion.  Spietliolf ''^  found 
that  pure  caseous  material  is  usually  free  from  even  traces  of  albumose 
and  peptone,  but  the  caseous  material  at  the  periphery  mixed  with 
tissue  elements  contains  them  in  very  small  quantities,  suggesting  that 
at  the  periphery  of  caseous  areas  some  slight  autolysis  does  occur.  The 
fact  that  B.  tuhcrculosis  is,  itself,  very  poor  in  proteolytic  enzymes  as 
compared  with  most  other  bacteria  may  be  another  factor.  When 
leucocytes  are  attracted  into  a  tuberculous  focus  softening  goes 
on  rapidly,  showing  that  there  is  no  loss  of  digestibility  of  the  caseous 
material,  but  merely  a  lack  of  enzymes.  Pus  from  a  cold  tuberculous 
abscess  will  not  digest  fibrin,  but  if  iodoform  is  injected,  leucocytes 
enter  in  great  numbers,  softening  is  rapid,  and  the  pus  will  then  di- 
gest fibrin  (Heile^®).  On  serum  plates  tuberculous  pus  produces  no 
digestion  unless  a  secondaiy  infection  or  other  cause  has  resulted  in 
a  local  accumulation  of  leucocytes.^  Tuberculous  material  contains, 
like  the  lymphocytes,  an  enzyme  which  is  proteoh'tic  in  acid  media 
and  which  is  inhibited  by  normal  serum  (Opie  and  Barker-). 

Correlation  of  Histological  and  Chemical  Changes. — A  careful  study 
of  the  relationship  of  the  chemical  changes  produced  by  autolysis,  to 
the  histological  changes  of  necrosis  and  autoh'sis,  has  been  made  by 
H.  J.  Corper,^^  and  colored  plates  published  together  with  analytical 
figures  make  it  possible  to  correlate  at  a  glance  the  structural  and 
chemical  changes  of  necrobiosis.  Corper  found  that  in  the  early 
stages,  characterized  by  a  high  grade  of  pycnosis  but  no  further  nu- 
clear changes,  the  nucleins  are  still  intact;  but  with  well  developed 
karyorrhexis  and  beginning  karyolysis,  some  ten  per  cent,  of  the  nu- 
clein  nitrogen  has  become  soluble  in  the  form  of  purine  bases.  Wlien 
karyolysis  is  completed  so  that  no  more  nuclei  remain  in  a  stainable 
condition,  only  twenty-eight  per  cent,  of  the  purines  was  found  to 
have  been  decomposed  to  free  purine  bases,®^'*  the  remaining  seventy- 
two  per  cent,  being  intact  although  unstainable.  This  rather  surpris- 
ing observation  indicates  that  the  stainable  chromatin  represents  but 
about  one-fourth  of  the  nucleins  of  the  cell,  which  is  in  accord  with 
the  views  of  Hammarsten  and  others.  The  lecithin  disintegrates 
somewhat  more  completely,  about  one-half  or  two-thirds  being  disin- 
tegrated by  the  time  nuclear  destmetion  is  complete,  after  which  this 
and  all  other  autolytic  change  is  slow.     The  change  from  coagulable 

97a  .Jour.  Exp.  Med.,  1914    (19),  38-3. 
05  Cent.  f.  inn.  Med.,  1904   (25),  481. 
90  Zeit.  klin.  IMed.,  1904    (55),  508. 
1  See  Wiens,  Joe.  citJ5 
2. Jour.  Exper.  Med.,   1900    (11),  686. 
93  Jour.  Exper.  Med.,   1912    (15),  429. 

93a  Marshall  (Jour.  Biol.  Chem.,  1913  (15),  81)  has  also  found  that  much  of 
the  nucleic  acid  remains  unaltered  in  autolysis  of  thymus. 


100  j:\z)ui:s 

to  non-coajrulahle  forms  of  nitroficn  was  as  follows:  Normal  spleen, 
noii-coagulable  iiitrojren,  5.7  per  cent,  of  the  total;  sta^e  of  marked 
pycnosis,  without  rhexis  or  lysis,  7.4  per  cent.;  stage  of  karyorrhexis 
and  early  karyolysis,  26.5  per  cent.;  stage  of  complete  karyolysis,  30.3 
per  cent.  That  is,  when  nuclear  structures  in  the  spleen  have  lost 
their  staining  properties  entirely  througli  autolysis,  about  72  per  cent, 
of  the  nuclein  nitrogen,  50  per  cent,  of  the  insoluljle  i)hosphorus  com- 
pounds, and  70  i)er  cent,  of  the  coagulnlilc  nitrogen,  are  .still  intact, 
and  about  two-tliii-ds  of  the  lecitliiu  remains  in  comijlex  organic  com- 
binatif)ns. 

Liver  Degenerations, — The  relation  of  tlie  disintegration  observed 
in  phosphorus-poisoiiiiKj  and  acutf  i/dloir  alrophu  to  the  experimental 
autolysis  of  the  liver  has  been  tlie  objeet  of  much  study.  Salkowski 
origiiuilly  pointed  out  that  the  same  jn-oducts  were  found  in  the  blood, 
urine,  and  liver  tissue  in  acute  yellow  atrophy  as  are  produced  in 
autolysis.  Jacoby -^  found  that  the  livers  of  dogs,  taken  just  as  the 
animals  Avere  dying  of  phosphorus-poisoning,  contained  free  leucine 
and  tyrosine;  also,  he  found  that  the  rate  of  autolysis  of  such  livers 
after  removal  from  the  body  was  much  greater  than  in  normal  livers. 
The  oxidizing  ferments  (aldehydase)  are  not  destroyed  by  the  proc- 
ess. He  found  that  addition  of  minute  amounts  of  phosphorus  to 
liver  enzymes  did  not  increase  their  proteolytic  power;  nevertheless, 
he  seems  inclined  to  assume  that  in  phosphorus-poisoning  alteration 
in  the  autolytic  "enzymes  is  an  important  factor  in  the  liver  degen- 
eration. It  would  seem  much  more  probable  that  phosphorus  is  a 
poison  that  kills  cells  and  does  not  destroy  their  antolytic  enzymes, 
hence  favoring  autolysis.  The  liver  degeneration  following  chloro- 
form poisoning  may,  perhaps,  be  explained  in  a  similar  way,  the  cells 
behaving  exactly  as  bacteria  would  do  under  the  same  conditions. 
Taylor  *  has  analyzed  several  livers  in  degenerative  conditions  for 
amino-acids  and  found  them  only  in  one  liver,  which  showed  necrosis 
])robably  due  to  chloroform  poisoning,  ami  wliich  was  from  a  case 
clinically  resembling  acute  yellow  atrophy.  Here  he  obtained  4  gm. 
of  leucine,  2.2  gm.  of  tyrosine,  and  2.3  gm.  of  arginine  nitrate. 
AValdvogel  and  Tintcmann,"'  in  phosphorus  livers,  found  an  increase 
in  ])i-()tagon,  jecoi'in,  fatty  aeids,  cholesterol,  and  neutral  fat.  while 
lecithin  was  decreased.  Wakeman  "  found  arginine,  histidine,  and  ly- 
sine decreased  in  jiliospliorus  livei's  in  pi'oj)oi1  ion  to  the  total  nitro- 
<ren,  indicating  that  the  ])i'otein-spli1t  ing  enzyme  in  this  condition 
eitlu'r  i)icks  out  certain  varieties  of  pi'oteins  lirst,  or  removes  the 
nitrogen-rich  constituents  most  rapidly.' 

•■i  Z«-it.  f.  plivsiol.  Cli.'iM..   l!i(l()   I  lid  I.   171. 
4  Univ.  of  ("'alif.   I'lililic.    (  patliol.) ,   1!)(»4    (1).  4:i. 
oCent.  f.  Path.,   l!t()4    (15).  <)7. 
n  Ik'il.  kliii.   Wocli..   l!t()4    (41),   lOtn. 

7  ( 'on^idcialilc  tjiiant  it  ics  of  ainiiio-aciils  of  \arioiis  sorts  liavi'  lu'cii  isolattnl 
from   llii'   li\rr   in  ariilc  yellow  atropliy   and   chlorofoiiii   iiccidsi:    liy    Wolls    (.lo\ir. 


AUTOLY.'^IS  J\   I'ATIIOLOdlcA'    /V.'Ov'FA^^.S'  lOl 

It  is  probable  that  many  poisons  may  destroy  the  liver  cells  to 
snch  an  extent  that  they  cannot  maintain  their  normal  chemical 
('(luilibriuni,  witlinut.  at  the  same  time,  destroying  the  autolytic  eu- 
/yiiu's.  When  this  oceurs,  tlie  liver  nndergroes  antolysis,  and  we  f;et 
marked  degenerative  changes  Avith  appearance  of  amino-acids  in  the 
blood  and  urine,  reduction  in  coagulability  of  the  blood  and  numer- 
ous hemorrhages,  giving  a  picture  both  clinically  and  anatomically 
more  or  less  like  that  of  typical  acute  yellow  atrophy.  Chloroform 
is  a  poison  that  stops  cell  activities  Avithout  destroying  the  proteolytic 
enzymes,  hence  the  cells  undergo  autolysis,  and,  as  a  result,  we  have 
many  eases  of  what  appears  to  be  aeiite  yellow  atrophy  following 
chloroform  anesthesia.  The  liberation  of  IICl  in  the  liver  cells  dur- 
ing chloroform  poisoning,  as  demonstrated  by  Evarts  Graham,"''  may 
be  largely  responsible  for  the  rapid  disintegration  of  the  liver  in  this 
condition. '''  (See  "Acute  Yellow  Atrophy,"  Chap,  xviii.)  Probably 
the  liver  changes  in  puerperal  eclampsia,  and  in  streptococcus  and 
other  septicemias  are  of  a  similar  nature.^  Autolysis  of  fatty 
livers  in  tuberculosis  is  said  to  yield  more  lactic  acid  than  the  livers 
from  other  conditions  (Yousscmf),  suggesting  defective  oxidative 
powers. 

Postmortem  changes  are  undoubtedly  due  to  two  factors,  bacterial 
action  and  autolysis.  In  tissues  kept  at  a  low  enough  temperature 
to  exclude  bacterial  action,  but  not  so  low  as  absolutely  to  stop  enzyme 
action,  there  occurs  a  slow  autolysis;  this  constitutes  the  "ripening" 
process  of  meat.  Fish  flesh  may  also  ripen  when  made  sterile  in 
saturated  salt  solutions,  as  Schmidt-Nielsen  **  has  shown  occurs  with 
salted  herrings ;  oxy-acids  and  xanthine  bases  being  prominent  among 
the  products.  The  softening  of  muscles  in  rigor  mortis  is  probably 
also  an  autolytic  manifestation,  as  muscles  contain  proteases  acting 
best  in  acid  medium,  and  the  muscle  is  known  to  become  increasingly 
acid  after  circulation  ceases  within  it.  The  short  duration  of  rigor 
mortis  when  the  body  is  kept  warm,  and  its  early  disappearance  when 
death  has  been  preceded  by  muscular  exhaustion  (which  increases  the 
acidity),  agree  with  this  view.  The  early  postmortem  softening  of 
many  organs  in  pathological  conditions  is  also  probably  an  autolytic 
manifestation.  Flexner  ''*  has  called  attention  to  this  in  relation  to 
the  softening  of  the  parenchymatous  organs  in  acute  infectious  dis- 

Exper.  Med.,  1907  (9),  027;  Jour.  Biol.  Chem.,  1908  (5),  129)  :  Imt  tlic  value  of 
these  figures  is  questionable  bwause  it  is  possible  that  the  alcohol  in  wliicii  the 
tissues  were  kept  before  analysis  Avas  not  strong  enough  entirely  to  prevent 
autolysis   (Wells  and  Caldwell,  Jour.  Biol.  Chem.,  1914    (19),  57). 

"a  Jour.  Exp.  :Med.,   191,t    (22),  48. 

ThQuinan  (Jour.  Med.  Res.,  1915  (32),  73)  found  no  change  in  the  rate  of 
in  ritro  autolysis  of  liver  tissue  from  experimental  chloroform  poisoning.  It  was 
found  increased  bv  phlorhizin  ( Satta  and  Fasiani,  Arch,  di  Fisiol.,  1913  (11). 
391). 

swells,  Jour.  Amer.  Med.  Assoc,  190G   (40),  341. 

9  Hofmeister's  Beitrjige,  1903    (3),  207. 


102  ENZYMES 

eases,  such  as  typhoid  and  septicemia.  Schumni  noted  great  auto- 
lytic  activity  in  a  swollen  spleen  from  a  case  of  perityphlitis. 

Histological  chajigos  are  produced  by  autolysis  in  the  organs  after 
death  that  are,  as  might  be  expected,  much  like  those  seen  in  necrotic 
areas.^^  At  first  the  changes  resemble  those  of  parenchymatous  de- 
generation (cloudy  swelling),  and  often  there  is  an  apparent  increase 
in  fat.  which  is  probably  due  to  liberation  of  masked  fat  through 
the  destruction  of  the  protein.^-  Nuclear  staining  is  lost  (karyolysis), 
and  eventually  even  cell  forms  become  indistinguishable,  but  this  does 
not  ordinarily  become  comi)lete  in  autolysis  without  bacterial  com- 
plication.     (See  p.  1)9  on  chcmiral  changes  of  postmortem  autolysis.) 

Still-born  children  that  have  been  carried  for  some  time  after  death 
usually  show  considerable  disintegration  of  the  viscera,  especially  the 
liver.  This  is  undoubtedly  due  to  autolysis,  which  Schlesinger  ^^  has 
shown  can  begin  before  birth  if  the  fetus  dies  /;(  utcro. 

Autolysis  in  Relation  to  Infection. — According  to  Conradi  ^*  the 
substances  produced  in  tissue  autolysis  have  a  decided  inhibiting  effect 
upon  bacteria,  which  apparently  depends  upon  the  antiseptic  proper- 
ties of  the  aromatic  derivatives  that  are  split  out  of  the  protein  mole- 
cule in  autolysis.  This  action  is  manifested  not  only  in  vitro,  but  the 
autolytic  products  will  also  render  harmless  lethal  doses  of  certain 
bacteria  if  they  are  injected  simultaneously  with  the  bacteria  into  an 
animal.  One  specific  class  of  products  of  autolysis  which  is  strongly 
bactericidal  is  the  soaps.^^  It  may  well  be  questioned,  however, 
whether  enough  of  these  substances  ever  accumulates  in  infected 
tissues  during  intra  vitam  autolysis  to  have  much  effect  upon  the  in- 
fecting bacteria;  yet  this  property  may  possibly  explain  the  steriliza- 
tion of  old  pus  collections  and  similar  infected  accumulations  within 
the  body.  The  bacteria  themselves  also  produce  autolytic  products 
that  are  powerfully  bactericidal.      (See  "Bacteria,"  Chap,  iv.) 

Blum  ^^  found  that  the  autolytic  products  of  lymph-glands  neu- 
tralized tetanus  toxin,  but  were  inactive  against  diphtheria  toxin  and 
cobra  venom.  Products  from  other  autolyzed  organs  and  from  fresh 
lymph-glands  were  without  influence  on  the  tetanus  toxin.  The  anti- 
toxic principles  of  the  autolytic  product  were  destroyed  by  heating, 
weakened  by  acids  and  alkalies,  and  in  other  respects  showed  prop- 
erties strikingly  like  those  of  true  antitoxin.  It  is  quite  possible  that 
bacterial  toxins  may  be  destroyed  by  autolytic  enzymes,  for  Baldwin 

11  More  fully  discussed  liy  Wells.  .Tour.  Mod.  Kesearcli,  1006   (15).  149. 

laSit'frert  ( llofmeistor's  Beitr.,  1901  (1),  114)  found  no  actual  increase  in  fats 
and  fatty  acids  in  atifolysia  even  wlien  an  increase  was  api)arent  histolopicallv, 
altliou;rli  ether-solnlile  materials  of  other  nature  than  fat  may  be  increased.  See 
als()  11  ess  and  Sa\l.  Xircliow's  Arch.,  1910   (202),  149. 

13  llofmeister's  Reitr.,  1903   (4),  87. 

1*  IlofmeisU'r's  Heitr.,  1901    (1),  19.'?.     See  also  Bilancioni  'Wn  and  Alnui'na  srb 

15  Se<'  Lamar,  Jour.  Kxp.  Jled.,  1911    (13),  1 

lollofmeister's  Heitr.,  1904    '5i,   142. 


LEUKEMIA  103 

and  Levene  ^'  have  shown  that  trypsin,  pepsin,  and  papain  destroy 
tetanus  and  diplitheria  toxin,  while  tuberculin  is  destroyed  by  trypsin, 
but  not  readily  by  pepsin,  possibly  because  it  is  of  a  nucleoprotein 
nature.  The  leueocytic  proteases,  however,  seem  not  to  attack  either 
toxins  or  living  bacteria  (Jochmann).  Bertolini '^  states  that  auto- 
lyzing:  liver  will  destroy  diphtheria  toxin. 

On  the  other  hand,  there  are  many  pathogenic  bacteria  which  do 
not  secrete  their  toxic  materials,  but  store  them  up  within  the  cell 
body,  e.  g.,  typhoid,  cholera,  and,  indeed,  the  majority  of  pathogenic 
forms.  These  endotoxins  are  probably  liberated  from  the  bacteria 
only  through  digestion  of  their  cells,  either  by  their  own  autolytic 
enzymes  or  by  the  enzymes  of  the  infected  tissues  and  leucocytes. 

Leukemia. — The  al)undant  elimination  of  uric  acid  and  other  pu- 
rine bodies  in  the  urine  in  leukemia  testifies  to  the  great  amount  of 
destruction  of  nucleoprotein  that  is  going  on  during  the  disease,  and 
these  are  probably  derived  from  the  autolysis  of  leucocytes,  which  per- 
haps depends  on  the  relatively  large  proportion  of  leucocytes  to 
serum.  Schumm  ^^  has  found  that  leukemic  spleens  and  bone  mar- 
row autolyze  rapidly  and  completely,  and  he  isolated  many  of  the 
cleavage  products  of  protein  digestion  from  such  autolysates. 

Leucocytes  from  myeloid  leukemia  liquefy  alkaline  gelatin  vigor- 
ously, but  those  from  lymphatic  leukemia  do  not;  the  liquefaction  is 
inhibited  by  normal  serum  (Stern  and  Eppenstein).-''  By  the  serum 
plate  method  this  observation  has  been  much  extended,  and  the  hetero- 
l>i:ic  action  of  the  lecocytes  has  been  found  limited  to  the  neutro- 
phile  granules.  In  neutral  media  evidence  is  obtained  of  the  presence 
of  protease  in  the  lymphoc}i:es  of  chronic  lymphatic  leukemia  and  the 
leucocytes  of  acute  and  chronic  myeloid  leukemia ;  maltase,  lipase  and 
amylase  are  found  in  both  types  of  cells,  and  oxidase  in  the  granular 
cells  derived  from  the  marrow  (^Morris  and  Boggs).-^  v.  Jaksch." 
Erben.-^  and  others  have  noted  the  occurrence  of  peptones  and  albu- 
moses  in  leukemic  blood,  particularly  if  removed  postmortem.  The 
improvement  in  leukemia  that  follows  j:--ray  treatment  is  associated 
with  an  increased  nitrogen  elimination,  probably  due  to  autolysis  of 
disintegrating  cells,-*  although  j--rays  have  no  appreciable  effect  upon 
the  leueocytic  proteases  in  vitro  (^Miiller  and  Jochmann).  (See  also 
"Leukemia,"  Chap,  xi.) 

IT  Jour.  Med.  Research,  1001    (6),  120. 
isBiochem.  Zeit.,  1913   (48),  448. 

19  Hofmeisters  Beitr.,  1003    (.3),  576:    1905    (7).  175. 

20  See  discussion  of  leueocytic  enzymes,  p.  94.  Longcope  and  Donhauser  (.Jour. 
Exper.  !Med.,  1908  (10),  618)  found  proteases  in  the  large  lymphocytes  in  acute 
leukemia,  which  were  most  active  in  an  alkaline  medium. 

21  Arch.  Int.  :\red.,  1911    (8),  806. 

22  Zeit.  f.  phvsiol.  Chem..  1892   (16).  243. 

23  Zeit.  f.  klin.  Med.,  1900  (40),  282;  Zeit.  f.  Heilkunde.  1903  (24),  70:  Hof- 
meister's  Beitr..  1904   (5).  461. 

24Mus5er  and  Edsall.  Univ.  Penn.  Med.  Bull.,  1905    (18),  174. 


104  i:\/.yMiJs 

Tumors. — l^i-obably  because  of  tlie  jiTeat  amount  of  necrosis  that 
is  constantly  <i-oin^-  on  in  all  nialiji-nant  jiTowths,  with  subsequent  di- 
gestion of  the  dead  cells,  autolytic  products  are  present  in  them  in 
very  considerable  amounts.  This  was  first  demonstrated  by  Petry,-'" 
who  found  that  carcinomata  of  the  breast  contained  much  of  their 
nitrogen  in  c()m])ounds  not  coagulated  by  heat,  while  in  the  normal 
eland  practically  all  is  coagulable.  Tie  also  demonstrated  an  autolytic 
property  in  tumor  tissue,  showing  that  tumor  cells  do  not  ditt'er  in  this 
respect  from  normal  cells.  l-Jeebe -"  found  i)i'()(lucts  of  autolysis  con- 
stantly present  in  several  tumors;  namely,  a  carcinoma  of  the  broad 
ligament,  a  hypernephroma,  an  angiosarcoma,  and  a  round-cell  sar- 
coma. 

Neuberg -'  found  that  while,  according  to  other  observers,  most 
enzymes,  as  well  as  bacteria,  are  very  susceptible  to  the  action  of 
radium  rays,  the  autolytic  enzymes  of  cancer  cells  are  an  exception, 
for  cancer  tissue  exposed  to  radium  undergoes  autolysis  much  faster 
than  cancer  tissue  not  exposed  to  radium ;  .^-rays  are  less  active  in 
this  respect.  He  attributes  the  effects  of  radium  on  cancer  to  its 
deleterious  effects  on  the  oxidizing  and  other  enzymes  of  the  cells, 
destroying  their  activities,  which  results  in  destruction  of  the  cells 
by  the  autolytic  enzymes.--  A  cancer  of  the  stomach  was  found  to 
contain  autolytic  enzymes  capable  of  digesting  lung  tissue  (pepsin 
was  excluded)  and  autolyzed  cancers  yielded  much  pentose.  Blu- 
menthal  and  Wolf  -"  believe  that  tumor  tissues  have  particularly  active 
autolytic  enzymes,  since  liver  tissue  added  to  tumor  tissue  underwent 
autolysis  much  more  rapidly  than  normal ;  but  tumors  do  not  cause 
digestion  of  serum  plates  unless  many  leucocytes  are  present  (IMiiller 
and  Kolaczek).^^  Cancer  extracts  digest  peptids  in  ways  different 
from  normal  tissues,  which  seems  to  indicate  some  fundamental  ab- 
normality in  their  metabolism  (Abderhalden,-"  Neuberg''-).  The  al- 
most constant  presence  in  gastric  juice  of  patients  with  carcinoma 
of  the  stomach,  of  ereptases  hydrolyzing  proteoses  and  peptids,  is 
generally  attril)ut('d  to  the  disintegration  of  the  cancer  with  libera- 

2o  Zeit.  f.  plivsiol.  Choni.,  1899    (27),  398;  Hofmoister's  Beitr.,   1!)()2    (2).  04. 

2<i  Amer.  Joiir.  Pliysiol.,   1904    (11),   139. 

27  Zeit.  f.  Krc'l)sforscl)uii<j:,  1904  (2),  171;  P.orlin.  klin.  Woch.,  1904  (41), 
1081:  ibid.,  1905    (42),  118;  Arb.  Path.  Inst.  Berlin.  190(1.  p.  o93. 

-•«  Wohlfrcnnitli,  Borl.  klin.  Woeh.,  1904  (41).  704,  found  that  autolysis  in 
tuberculous  luiifr  tissue  was  three  or  four  times  more  rapid  wlieu  exposed  to 
radium  rays,  lleile  (Arch.  klin.  Cliir.,  lOOo  (77).  107  I  looks  ujioii  the  favorable 
etlects  of  a'-rays  as  partly  jiroduced  by  their  liberation  of  autolytic  eii/ymes 
from  the  loucocvtcs. 

23  Med.   Klini'k.,   1905    (1),  No.  7. 

■•'•MiilJer  and  Kolaczek,  :Miincli.  nied.  Wodi..  1907  ( .")4 ) .  3.")4 :  lless  and  Saxl. 
Wien.    klin.    Woch.,    l!l(ts    (21),    1183;    l\c]>ino\v.    Zeit.    f.    K rebsforscli..    I'.Ki'.i    (7i. 

r)i7. 

31  Zeit.  plixsiol.  CIm'iii.,   1910    ((>(i),  277. 
s2Bio<  licin'.   /cii.,    1 1)  1(1    (2(i),  344. 


T!  Moh'X  105 

tioii  of  these  enz.ymes.''''  Tumors  also  contain  nuclease^*  to  disinte- 
{^rate  their  nucleic  acid,  and  the  same  outfit  of  purine-splitting  en- 
zymes as  normal  tissues,-'"'  so  that  in  rep:ard  to  the  nucleoproteins  of 
tumors  autolysis  follows  the  same  course  as  in  normal  tissues. 

The  non-cancerous  livers  of  cancerous  patients  were  found  by  Yous- 
souf  •'"  to  jiroduce  more  lactic  acid  duringr  antiseptic  autolysis  than 
did  livers  in  other  conditions.  Autolysis  of  orfjans  of  cancer  patients 
is  about  as  rapid  as  normal  ( Cohvell "' ).  Several  observations  have 
sugrp-ested  that  tumor  tissues  mig-ht  contain  ])r()teolytic  enzymes  dif- 
fering from  those  of  normal  tissues  especially  in  their  ability  to  di- 
gest heterologous  normal  tissues,  ])ut  at  present  this  work  needs  con- 
firmation and  amplification  before  it  can  carry  the  weight  of  specula- 
tion which  has  been  heaped  upon  it.''- 

^licheli  and  Donati  ■■'  attribute  the  hemolytic  properties  possessed 
by  extracts  of  malignant  tumors  to  the  products  of  autolysis  that  are 
present,  which  Petry  has  also  demonstrated  to  produce  hemolysis. 
Emerson  *°  attributes  the  disappearance  of  HCl  from  the  gastric 
juice  in  carcinoma  of  the  stomach  to  neutralization  by  basic  products 
of  autolysis,  a  hypothesis  that  nmy  well  be  questioned.  (See  also 
"Tumors,"  Chap,  xvii.) 

Varioiis  other  intracellular  enzymes  have  been  described,  -wliich  for  the  most 
part  iiave  as  yet  no  significanee  in  pathology.  An  exception  is  fiJn-ln  fcnnent, 
which  will  be  considered  fully  in  discussing  thrombosis.  Ferments  coagulating 
milk  seem  to  be  widely  spread  in  the  tissues.  The  precipitation  of  plastein  from 
proteose  solution  by  organ  extracts  (Xiirnberg)  may  be  either  the  effect  of  a 
coagulating  ferment  or  due  to  reverse  action  of  the  proteases.  Ferments  s[)lit- 
ting  specifically  maltose,  lactose,  sucrose,  glucosides.  and  nucleoproteins  have 
been  described,  and  the  glycogenolytic  ferment  is  probably  nearly  universallv  pres- 
ent. Other  enzymes  decomposing  amino-acids  into  ammonium  compounds  may 
also  exist.  The  enzymes  acting  specifically  upon  the  nucleic  acids  and  the  ])urine 
bodies  are  discussed  in  Chapter  xxi. 

33  See  Jacques  and  ^Yoodyatt,  Arch.  Int.  :\Ied.,  1012  (10),  .lOO:  Ilamlmrger, 
Jour.  Amer.  Med.  Assoc.  1912   (59),  847. 

34  Goodman,  Jour.  Exp.  ^^led..  1912    (15),  477. 

33  Wells  and  Long,  Zeit.  Krebforsch.,  1913    (12),  .598. 
36Virchow's  Arch.,   1912    (207),  .374. 

37  Arch.  Middlesex  ITosp..  1910    (19).  55. 

38  See,  for  example,  Rulf,  Zeit.  Krebsforsch.,  1906  (4).  417:  Miillcr,  Cent.  inn. 
Med.,  1909   (30),  89. 

3oRiforma  med..  1903    (19).   1037. 

40  Deut.  Arcli.  klin.  Med.,  1902   (72),  415. 


CHAPTER    IV 

THE  CHEMISTRY  OF  BACTERIA  AND  THEIR 
PRODUCTS 

STRUCTURE  AND  PHYSICAL  PROPERTIES  ' 

In  structure,  as  in  uearly  all  other  respects,  bacterial  cells  stand 
inteiinediate  between  the  cells  of  ordinary  plant  and  animal  tissues. 
Their  cell  wall  seems  to  be  generally  more  hig-hl}-  developed  than  that 
of  animal  cells,  and  less  so  than  the  wall  of  most  plant  cells.  In 
composition,  however,  the  wall  is  more  closely  related  to  animal  than 
to  vegetable  tissues.  The  much  vexed  question  as  to  the  existence 
or  non-existence  of  a  nucleus  seems  to  be  best  answered  by  Zettnow, 
who  considers  that  the  portion  of  the  bacterial  cell  usually  made 
evident  by  ordinary  staining  methods  consists  of  a  mixture  of  nuclear 
substance  {chroiiiati)i)  with  non-chromatic  substance  {endoplasm)  ; 
the  outer  membrane,  which  requires  special  methods  for  its  satisfac- 
tory demonstration,  consists  of  a  modified  cytoplasm  {ectoplasm). 
Some  bacteria  consist  chiefly  of  chromatin  (e.  g.,  vibrios),  but  the 
proportion  of  the  ditit'erent  elements  varies  greatly,  not  onh^  in  dif- 
ferent varieties,  but  also  in  the  same  variety  under  ditferent  con- 
ditions. The  fact  that  the  chromatin  is  not  aggregated  into  the  usual 
nuclear  form  may  be  ascribed  to  the  low  stage  of  development  reached 
by  bacteria  in  the  scale  of  evolution ;  or,  as  Vejdovosky^  has  suggested, 
to  the  extremely  rapid  rate  of  cell  division  in  the  bacteria  which  pre- 
vents the  chromatin  from  appearing  in  the  resting  stage  whicli  a 
nucleus  constitutes.  Finer  structures  witliiu  tlie  bacterial  cell  have 
as  3'et  been  onl}^  imperfectly  discerned. 

The  thickness  of  the  ectoplasm  varies  greatly  even  in  the  same 
species,  being  generally  greatest  in  older  cultures.  In  some  forms 
the  ectoplasm  may  constitute  one-half  of  the  total  mass  of  the  cells. 
The  capsule  seems  to  arise  through  a  swelling  of  the  ectoplasm,  and 
is  probably  present  in  at  least  a  rudiniontary  stage  in  all  bacteria 

LMiguhi).' 

F^lasmolysis  and  Plasmoptysis. — I'mler  conditions  of  altered 
osmotic  pressure  the  bacterial  cell  behaves  quite  similarly  to  the  plant 

1  In  this  chapter  reforcnees  will  not  ponorally  he  <iiven  that  can  he  found  hy 
consul(in<j:  Kolio  and  \Vass(>iinann's  TIandliucli.  A  jxcncral  consideration  of  the 
Bio]o<,'y  of  the  Bacteria,  incliidinjj  refercn<es  1o  llie  ctVecls  of  lijilit,  heat,  osmotic 
pressure,  etc.,  is  friven  hv  Miiller,  Krgb.  der  Physiol.,  1004  (4),  13S;  concerninc 
their  chemistry  see  H.  Fisciier,  Lafar's  IIandl)uch  der  Teclinischcn  ^lykologie, 
1908    (1),  222." 

lOfl 


CHEMICAL  COMPOSITION  OF  BACTERIA  107 

cell.-  If  placed  suddenlj^  in  a  solution  of  higher  osmotic  pressure  than 
the  one  in  which  it  has  been,  the  cell  contents  shrink  away  from  the  cell 
wall  {plasmoli/sis)  indicating  that  there  exists  a  semipermeable  mem- 
brane through  which  water  passes  more  rapidly  than  salts.  If  the 
change  in  osmotic  pressure  is  gradual,  the  bacteria  accommodate 
themselves  to  it  by  the  slow  diffusion  of  the  salts  through  the  cell 
membrane,  indicating  that  it  is  not  absolutely  semipermeable.  Dif- 
ferent bacteria  behave  differently,  some  bacteria  not  l)eing  plasmolyzed 
by  solutions  that  plasmol^^ze  others.  As  a  nde,  old  bacteria  plas- 
mol^'ze  more  rapidly  than  young,  and  in  some  varieties  there  seems 
to  be  a  spontaneous  plasmolysis,  to  which  lias  been  attributed  the 
irregular  staining  of  diphtheria  and  tubercle  bacilli,  the  polar  stain- 
ing of  plague  bacilli,  etc.  Plasmolysis  occurs  only  in  living  bacilli, 
but  does  not  necessarily  cause  death.  The  Gram-staining  bacteria 
cannot  generally  be  plasraolyzed,  and  contain  more  water.^ 

AVlien  bacteria  pass  from  solutions  of  higher  osmotic  concentration 
into  solutions  of  lower  concentration,  tlie  phenomenon  of  2)^dsmoi)tysis 
is  produced.  The  cell  contents  swell  until  the  cell  wall  gives  way  at 
some  point,  and  then  exude  as  glistening  drops,  which  may  become 
detached  from  the  wall  and  escape  free  into  the  fluid.  Plasmoptysis 
is  shown  best  by  bacteria  that  have  been  grown  on  salt-rich  media 
before  being  placed  in  the  salt-free  fluid.  Not  all  varieties  of  bacteria 
can  be  made  to  undergo  this  change,  depending  probably  upon  the 
degree  of  permeability  of  their  cell  membranes  for  salts.  The  ex- 
posure of  the  naked  cell  contents  to  the  hypotonic  fluid  outside  the 
cells  makes  plasmoptysis  more  serious  for  bacterial  life  than  plasmo- 
lysis, but  how  often  either  process  plays  a  part  in  the  resistance  of 
infected  animals  against  bacteria  is  unknown. 

Chemotaxis. — Just  as  with  unicellular  animal  organisms,  bacteria 
respond  to  chemotaetic  influences,  in  general  being  attracted  by  sub- 
stances favorable  for  food,  such  as  peptone,  dilute  potassium  salts, 
etc.,  and  being  repelled  by  harmful  substances,  such  as  strong  acids 
and  alkalies.  Attempts  have  been  made  to  separate  different  organ- 
isms in  mixed  cultures  by  means  of  their  response  to  chemotaxis,  but 
without  striking  success.  It  is  possible  that  chemotaxis  may  play  a 
part  in  the  localization  of  bacteria  from  the  blood  stream  in  favorable 
localities,  just  as  leucocytes  are  attracted  to  points  of  injury,  but 
this  has  not  been  demonstrated.  (The  chemotaetic  influence  of  bac- 
teria upon  leucocytes  is  discussed  in  Chapter  x.) 

CHEMICAL  COMPOSITION 

This  varies  greatly,  not  only  between  different  species,  but  even 
in  the  same  species  grown  on  different  media ;  in  this  respect  bacteria 

2  Literature,  see  Gotschlich.  7\olle  and  \YasFerinann's  Handbuch,  vol.  1. 
sNicolle  and  Alilaire.  Ann.  Inst.  Pasteur,   1000    (23),  547. 


108  CHEMISTRY  OF  BAVTEIUA    AM)   Til  El  I!   I'RODUCTS 

are  much  more  modified  by  tlieir  enviroiinieiit  than  are  higher  or- 
ganisms. They  usually  contain  between  80  and  90  per  cent,  of  water. 
Grown  on  a  salt-rich  medium  they  yield  much  ash;  grown  on  a  pep- 
tone-rich medium  they  contain  nnich  protein ;  grown  on  a  fat-rich 
medium  they  contain  much  material  solubU^  in  ether.  Cholera  vibrios 
grown  on  a  bouillon  medium  contained  ()i).25  per  cent,  of  protein,  and 
25.87  per  cent,  of  ash,  whereas  the  same  organism  grown  on  Uschin- 
sky's  medium,  which  contains  no  proteins  but  only  various  simple 
chemical  compounds,  contained  but  35.75  ))er  cent,  of  protein  and 
13.7  per  cent,  of  ash  (Cramer).  Even  in  the  same  medium  two 
different  strains  of  the  same  organism  may  show  e(iually  great  dif- 
ferences: Two  strains  of  cholera  vibrios  grown  on  the  same  medium 
showed  respectively  65.63  per  cent,  and  34.37  per  cent,  of  protein. 
It  is  evident,  therefore,  that  quantitative  analyses  of  bacteria  show 
nothing  as  to  their  nature,  and  on  account  of  the  extreme  limits  of 
their  variation  ai'c  practically  valueless.  The  specific  gravity  of  bac- 
teria, generally  between  1.12  and  1.345,  also  varies  with  media  and 
age.^     In  an  electric  field  they  move  towards  the  anode."' 

Qualitatively  the  variations  are  not  so  great — all  bacteria  contain 
proteins,  lipoid  substances,  and  salts,  of  which  phosphates  are  most 
prominent  in  the  ash.  The  character  of  the  proteins  and  fats  of 
bacteria  grown  on  peptone  bouillon  is  (juite  the  same  as  when  they 
are  grown  on  protein-free  media.'"'^'  The  older  analyses  of  bacterial 
constituents  are  of  little  value.  Recent  studies  prove  that  the  chief 
constituent  of  the  cell  .contents  is  a  true  nucleoprotein  (Iwanoff  '^)  con- 
taining some  sulphur  and  iron ;  probably  many  of  the  "pyogenetic  pro- 
teins," "bacterial  toxalbumins, "  "bacterial  caseins"  of  earlier  investi- 
gators are  true  nucleoproteins.^^  The  stainable  substance  of  anthrax 
bacilli  behaves  as  if  it  were  a  chromatin,  while  the  spores  resemble  linin 
fRuzicka').  The  predominance  of  nuclein  compounds  is  shown  by 
Ruppel's  summary  of  the  composition  of  dried  tubercle  bacilli, 
namely,  in  per  cent.,  tuberculonucleic  acid,  8.5 ;  nucleo-protamine,  24.5  ; 
nucleo-protein,  26.5 ;  fatty  matter,  26.5 ;  inorganic,  9.2 ;  insoluble 
"proteinoid"  residue,  8.3.  In  a  water  bacillus  Nishimura  found 
xanthine,  guanine,  and  adenine,  indicating  the  presence  of  nucleo- 
protein; others  have  found  that  bacterial  nucleoprotcins  s])lit  off 
pentoses,  as  do  the  nucleoprotcins  of  higher  cells.  If  it  is  true  tliat 
bacterial  luiclco-proteins  contain  pentos(>  it  ranks  tluMU  with  tlie  plant 

4Rtip('ll,  Cent.  f.   I?akt.,   1007    (4.3),  4S7. 

•'"'  Jinx  ton,  Zcit.  ]»liysil<al.  C'licni.,  liKU;  (57),  47;  concprninfj  the  rlcctriral  cnn- 
ductirH)/  of  liactcria  sec  'Iliornton.  Troc.  T\oval  Soc,  London,  Sec.  B.,  1013  (85), 
3.31. 

•laTannira.  Zcil.  iilivsiol.  Clicni..   lOl.S    (SS),   lon. 

0  Hofnieistcr's  Heitr.,  1002  (1),  524  ;  liililio,-ra|.liy  by  Lus(i<:-.  Kollc  and  Wassor- 
mann's  Handliucli,  1013    (ii),  1302. 

«"  The  ])nri1v  of  inanv  of  llic  jJi-cparalions  \Mirk(<l  with  as  bacterial  niulcopro- 
teins,  is  vcrv  donlitful.   "  (See  W.-lis,  Zeit.  Inununiliit ..   lOl.'i   (19),  500.) 

7  Arch.    KnlwicklniiKsink..    I'KX;    (21).   30(i. 


CUHMIi'M.   CUM I'OSI'I  li)\    or  li.MJTEUl.X  109 

iiucleo-proteins,  for  animal  mu-leie  acids  contain  hexosc.  On  tlie  otiier 
liand,  Levene  found  in  bacterial  nucleic  acid  tlie  ]nriinidines  thyniine 
and  uracil,  wliicli  are  respectively  characteristic  of  animal  and  ve<;e- 
table  nucleic  acids,  but  they  are  not  supposed  to  occur  in  botii.  Mary 
Leach  **  found  evidence  that  the  colon,  bacillus  is  largely  made  up  of 
nuclein  or  g-lyco-nucleoproteins,  but  contains  no  cclhdose.  Other  pro- 
teins, namely,  globulins  and  nucleo-albumins,  have  also  been  described 
as  constituents  of  the  bacterial  plasma. 

The  complete,  amino-acid  content  of  l)acterial  protein  does  not  seem 
to  have  been  worked  out.  althougii  the  workers  in  Vaughan's  labora- 
tory have  identified  many  of  the  usual  amino-acids  of  proteins  among 
tiie  products  of  hydrolysis  of  bacteria/'  Analysis  of  B.  inesmtericus 
shows  it  to  be  deficient  in  diamino-acids,  tyrosine,  glycocoll,  and  to 
contain  l(i.(i  per  cent,  of  glutamic  acid.^*^  Taraura  ^"-"^  found  i)lienyl- 
iilanine  and  valine  high  in  tubercle  bacilli  and  very  low  in  H.  diph- 
therup,  in  which  tyrosine  is  more  abundant.  In  an  azobaeterium, 
lysine  has  been  found  especially  abundant.^'"'  Cystine  has  been 
lacking  in  several  analyses.  Tamura  "'^  also  found  tluit  bacteria  can 
synthesize  from  simple  nonprotein  media  the  purines,  ])liosphatids  and 
the  typical  proteins  containing  the  aromatic  amino-acids.  This  syn- 
thetic activity  of  bacteria,  in  view  of  the  large  quantity  of  bacterial 
substance  in  feces,  may  possibly  be  of  importance  in  metabolism  studies, 
leading  to  erroneous  conclusions  as  to  utilization  or  synthesis  of  pro- 
teins by  the  subject.^"'' 

The  slimy  material  produced  in  cultures  by  some  varieties  of  bac- 
teria is,  at  least  for  certain  forms,  a  body  closely  related  to  or  identi- 
cal with  true  mucin, ^^  but  in  certain  cases  (B.  radicicola)  it  is  a  gum 
related  to  the  dextrans  and  free  from  nitrogen  (Buchanan).^-  Tu- 
bercle bacilli  grown  for  many  years  on  artificial  media  may  produce 
a  true  mucin  ( Weleminsky).^^  Ileim  ^^  considers  that  anthrax  bacilli 
also  produce  mucin.  Some  nonpathogenic  bacteria  contain  granules 
of  sulfur  in  their  protoplasm,  and  others  have  noteworthy  cjuantities  of 
iron  in  the  sheath. 

Bacterial  Carbohydrates. — The  earlier  descriptions  of  cellulose  or 
hemiccUulosc  in  the  cell  membrane  of  bacteria  have  been  contested. ^^^ 

8  Jour.  Biol.  Chem..  1906  (1).  46.3.  Full  bililioorapliy  on  Chemistry  of  Htu- 
teria.  See  also  Vaug-han,  "'Protein  Split  Products  in  Relation  to  Immunity  and 
Disease,"  Philadelphia,  1013. 

9  See  Wheeler,  Jour.  Biol.  Chem.,  1909   (6),  509. 

^0  Horowitz-Wlassowa,  Arch.  Sci.  Bioloitique,   1910    (15).  40. 

JoaZeit.  phvsiol.  Chem.,  1913    (87),  85;'  1914    (89),  289. 

lob  Omelianskv  and  Sieber,  Zeit.  phvsiol.  Chem.,  1913   (88),  445. 

lOcZeit.  phvsiol.  Chem.,   1913    (88)"    190. 

lod  Osborne  and  :\lendel.  Jour.  Biol.  Cheiu..    1913    (18).    177. 

11  Rettc;er,  Jour.  Med.  Research.  1903    (10),   101. 

12  Cent.  f.  Bakt.,  TI  Abt..  1909    (22).  371. 

13  Berl.  klin.  Woch.,   1912    (49),  1320. 
iiMiinch.  med.  Woch..  1904    (51),  426. 

"a  However,  Dreve.r    (Zeit.  ges.  Brauw.,  1913    (36),  201),  states  that  the  cell 


110  CHEMISTRY  OF  BACTERIA  ASD  THEIR  PRODUCTS 

Numerous  invosti<2:ators  have  reported  that  the  insoluble  bacterial  cell 
wall  consists  chiefly-  of  chitin,  which  on  being  split  with  acids  yields 
80  to  90  per  cent,  of  the  nitrogenous  carbohydrate,  glucosamin.^^  The 
distinction  is  a  very  important  one,  since  cellulose  is  a  typically  vege- 
table product,  while  chitin  is  eciually  typically  animal  in  origin,  being 
found  chiefly  in  the  shells  of  lobsters  and  crabs,  the  wings  and  cover- 
ings of  flies,  beetles,  etc.  Chitin  seems  to  be  a  polymeric  form  of 
glucose-amine,^'""  an  amino-carbohydrate,  just  as  cellulose  is  a  polymer 
of  a  simpler  carbohydrate.  Other  carbohydrates  seem  to  be  scanty  in 
the  bacterial  cell,  but  Tamura  ^^''^  does  not  accept  the  chitinous  nature 
of  bacterial  carbohydrate,  finding  in  tubercle  and  diphtheria  bacilli  a 
hemicellulose,  apparently  a  pentosan  yielding  1-arabinose  on  hydro- 
lysis. Wester  ^^"^  found  no  chitin  in  several  varieties  of  bacteria,  and 
cellulose  only  in  7>.  .ri/liuuDi ;  he  therefore  considers  it  probable  that 
bacterial  cell  walls  do  not  always  consist  of  the  same  substance.  Cramer 
could  find  no  glucose  in  any  variety,  although  there  are  some  bacteria 
that  contain  material  reacting  like  starch  with  iodin.  Levene,^*'  how- 
ever, found  in  B.  tuhcixulosis  a  substance  with  the  properties  of 
glyco<:'('ii. 

Bacterial  Fats. — By  staining  methods,  fats  have  been  recognized  in 
many  species,  and  by  extraction  with  fat  solvents  lecithin,  cholesterol, 
simple  fats,  and  specific  bacterial  fats  have  been  isolated ;  this  is  par- 
ticularly true  of  B.  tuberculosis,  which  owes  its  characteristic  staining 
])roperties  to  the  specific  fat-like  bodies  which  make  up  a  large  pro- 
portion of  its  entire  mass.^^  Numerous  studies  of  these  fats  of 
B.  tuherculosis  have  been  made  ^^  and  by  using  different  extractives, 
from  20  to  40  per  cent,  of  the  entire  weight  of  the  bacilli  has  been 
found  s()lul)le  in  fat  solvents.  Kresling  found  that  the  substance 
soluble  in  chloroform  had  the  following  composition: 

Free  fatty  acid 14. ."^R  per  cent. 

Neutral  fats  and  fatty  acid  osters 77.2.5     "  " 

Alcohols  obtained  from  fatty  acid  esters   ....  30.10     "  " 

Lecithin " O.lfi     " 

Substances  soluble  in   water 0.73     "  " 

Bulloch  and  ^Maclcod  found  that  ethereal  extracts  did  not  contain 
the  acid-fast  substance  which  they  consider  to  be  a  wax-like  alcohol, 
soluble  in  hot,  but  insoluble  in  cold  absolute  alcohol  or  in  ether. 
The  simple  fats  seem  to  be  formed  by  oleic,  isocetinic,  and  myristinic 

wall  of  veasts  contains  a  hemicellulose  and  a  nianno-dextran.  See  also  Kozniewski, 
Zeit.  phvsiol.  Chem..  1914    (00).  208. 

mSeoViehofer.  Ber.  Dent,  riiem.  fies..   1012    (301.  443. 

i-'i  Mor;rulis  states  that  chitin  consists  of  two  j)arts.  one  containincr  all  the 
glucose  and  amino  <:roups.  tlie  other  bcin?  a  stable  nitrogenous  compound  yielding 
no  plucoHC.      (Science.    l!»lf.    (44).  SCO.) 

isbZeit.   physiol.   Cliem..    1014    (.SO).   304. 

I'-cPharm.   Wcekbhid..    lOlC.    (.".3).    11S3. 

11  Jour.  Med.  Research.  1001    (0),  13r>. 

"Sec  Tamils  and  Pai.miez.  Compt.  Rend.  Soc.  Riol..   lOO.")    i,-)!)),  701. 

J 8  For  literature  see  Bulloch  iiiid  MachMKl.  .Tour,  of  lly-^i.-ne.  1004    (4).  1. 


CHEMICAL  COMPOSITIOX  OF  BACTERIA  111 

acids,  and  there  is  some  laiiric  acid  in  the  form  of  a  soap.  Cholesterol 
could  not  be  found  in  tubercle,  diphtheria  and  other  bacteria  examined 
by  Tamura,  although  there  probably  are  lipochroines  giving  the  cul- 
tures their  color.  There  is  still,  however,  much  disagreement  as  to 
whether  the  acid  fastness  of  tubercle  bacilli  depends  upon  waxes, 
alcohols,  fatty  acids,  or  lipoid-protein  compounds.^"  It  must  be  ad- 
mitted that  a  high  content  of  fatty  materials  is  regularly  present  in 
acid-fast  bacilli;  thus,  in  an  acid-fast  bacillus  isolated  from  leprous 
lesions,  34.7  per  cent,  of  fats,  fatty  acids  and  cholesterol,  and  1.7  per 
cent,  of  lecithin  were  found  by  Gurd  and  Denis.-" 

Tamura-"''  states  that  the  phosphatids  of  B.  tuberculosis  and  a 
saphrophyte  examined  by  him  were  not  lecithin  but  a  diaminophos- 
phatid,  although  diphtheria  bacilli  seemed  to  contain  lecithin.-"*^  He 
found  in  both  a  high  molecular  alcohol,  "mykol,"  to  which  he  ascribes 
acid-  and  Gram-fastness.  In  a  Gram-negative  bacillus  -°*^  he  found 
lecithin,  but  no  cholesterol  or  mykol.  By  growing  tubercle  bacilli 
on  suitable  media  they  can  be  made  to  lose  their  acid-fast  property, 
although  still  Gram-positive  (Wherry  ^'"^).  The  observation  of  ]\Iiss 
Sherman,-^  that  tubercle  bacilli  are  almost  absolutely  impenneable 
to  fat-soluble  dyes  which  stain  their  isolated  fats  well,  and  her  cor- 
roboration of  Benians'  demonstration  that  acid-fastness  depends  on 
the  integrity  of  the  bacillary  envelope,  make  the  role  of  the  fatty  sub- 
stances uncertain.  Their  high  content  in  unsaturated  fatty  acids 
gives  them  a  high  antitryptic  power  which  may  be  concerned  in  the 
defense  of  the  bacteria,  and  also  in  the  persistence  of  caseous  ma- 
terial in  tubercles  (Jobling  and  Petersen).-^-'' 

By  staining  with  Sudan  III,  Sata  --  demonstrated  fats,  not  only  in 
the  acid-fast  bacilli,  but  also  in  anthrax.  Staphylococcus  aureus,  B. 
mucosus,  and  actinomyces;  but  not  in  diphtheria,  pseudo-diphtheria, 
plague,  cholera,  and  chicken  cholera  bacilli,  or  in  members  of  the 
colon  group. -^  Only  a  few  bacteria  form  fat  on  agar  free  from 
glycerine,  but  potato  is  a  favorable  medium.  Ritchie  -*  obtained 
positive  fat  staining  in  B.  diphtheria'  and  anthracis,  but  not  in 
S.  pyogenes  aureus  or  M.  tetragenus,  although  these  last  forms  contain 

19  See  Camus  and  Pagniez,  Presse  MM.,  1007  (15),  65;  Devkc,  ^liinfh.  mod. 
Woch.,  1910   (57),  633. 

20  .Jour.  Exper.  :\[ed.,  1911    (14),  606. 
20aZeit.  phvsiol.  Chem..  1913   (87),  85. 
20hlhid.,   1914    (89),  289. 

20c  Ihid.,   1914    (90),  286. 

20d  Jour.  Infect.  Dis.,  1913   (13),  144. 

21  Jour.  Infect.  Dis.,  1913    (12),  249. 
2ia.Joiir.  Exp.  Med.,  1914    (19),  239. 

22  Cent.  f.  allg.  Path..  1900   (11).  97. 

23  Auclair  (Arch.  Med.  Exper..  1903  (15),  725)  contends  that  the  ether  and 
chloroform  extracts  of  many  patho>renic  bacteria  contain  important  toxic  sub- 
stances. Holmes  (CJuy's  TIosp.  Reports,  1905  (59),  155)  states  that  injection 
of  fattv  acids  from  tuljercle  bacilli  into  rabbits  causes  a  lymphocytosis. 

24  Jour.  Pathol,  and  Bact.,  1905   (10),  334. 


112  CHEMlSTh'Y  or  BAVTKltl  \    AM)   THEIR   I'UODLCTH 

cliomically  demonstrable  lipiiis.  Analyses  of  dififereiit  bacteria  show  a 
relatively  low  content  of  lipins  as  i-oiajjared  with  tubercle  bacilli,  vary- 
ing from  1.7  per  cent  in  B.  suhtilis  to  8.5  per  cent,  in  staphylococci 
(Jobling  and  Petersen).'-'-'  However,  the  degree  of  unsaturation  of 
the  fatty  acids  is  less  with  tuhei-clc  liacilli  than  with  other  bacteria 
examined  by  these  authoi's.  Extensive  studies  of  bacterial  fat  stains 
are  rej)()rted  by  Eiseiilxn'g,-''  but  ])ractically  nothing  is  known  of  the 
character  of  the  fally  or  lipoid  cDnslituonts  of  l)a('teria  outside  the 
acid-fast   grou]). 

Spores  diil'er  from  llicir  jiareiil  l)acteria  in  containing  a  nuich 
greater  proportion  of  the  solid  constituents  and  less  water.  In  molds 
Drymont  found  that  the  spores  contained  over  60  per  cent,  of  dry 
substance,  and  almost  all  the  water  was  so  held  as  to  resist  drying 
by  temperatures  below  boiling;  the  dry  substance  is  very  rich  in 
protein  and  poor  in  salts.  As  the  spores  may  lose  their  chromatin 
content  without  loss  of  capacity  to  propagate,  it  would  seem  that  this 
is  not  a  nuclear  chromatin  but  merely  a  reserve  food  supply.-""'^  The 
wall  of  the  spore  consists  of  a  "cellulose-like"  substance  and  a  very 
hygroscopic  extractive  matter.  The  great  resistance  of  spores  to  dry- 
ing and  to  heat  can  be  readily  understood  in  view  of  these  facts.  They 
contain,  and  perhaps  secrete,  active  enzymes  (Eifront).-'^  Flagella 
also  soom  to  be  composed  of  a  relatively  condensed  protein. 

Staining  Reactions. — The  staining  reactions  of  bacterial  cells  are 
much  as  if  the  bacteria  consisted  entirely  of  chronuitin,  so  that  at 
one  time  the  theory  prevailed  that  bacteria  consisted  merely  of  a 
nucleus  and  a  cell  wall,  without  any  true  cytoplasm.  The  demon- 
stration of  abundant  nucleoprotein  in  the  contents  of  bacterial  cells 
explains  their  staining  affinity  for  basic  anilin  dyes.  Owing  to  some 
unknown  differences  in  composition,  not  all  bacteria  are  stained 
equally  well  by  the  same  basic  dyes.  Although  the  staining  of  bac- 
teria depends  upon  a  chemical  reaction  between  the  nucleoproteins  and 
the  basic  dye,  yet  the  combination  is  not  usually  a  firm  one,  being 
readily  broken  by  weak  acids  in  most  cases.  That  the  decolorization 
of  bacteria  depends  u})oii  dissociation  of  the  dye-])roteiii  couiponnd 
is  shown  by  the  fact  that  absolutely  water-free  alcohol  will  not  de- 
colorize dry  bacteria,  nor  do  water-free  alcoholic  solutions  of  dyes 
stain  dehydrated  bacteria. 

Gram's  ^NFetiiod  of  staining  has  been  ascribed  to  llie  format  ion 
of  an  iodiii-])ararosanilin-protein  compound  which  is  not  easily  disso- 
ciated b\'  water'  in  the  case  of  bacteria  that  stain  b\"  tins  nu^thod,  and 
which  is  readily  dissociated  and  dissolved  out  in  the  case  of  bacteria 

24.1  .Tour.  Ivs]).  Mfd..  1!II4    CJO).  4.")(t. 
2-' Vircliow's  .Arcliiv.,  1!>1(I    (1!»!M.  ">n2. 
2-.a  Ruzifka,  ("cnt.  f.  l?al<t..  1014    (41).  (Ul. 
20Mon.  8c.  QuOTiu'villc.  1007,  p.  SI. 


BACTi:niAL  ENZY}fES  113 

that  do  not  retain  the  stain.  Only  pararosaniliu  dyes  (gentian  violet, 
methyl  violet,  victoria  blue)  form  such  combinations,  the  rosanilin 
dyes  not  beinji'  suitable. 

The  relation  of  bacterial  protein  to  Gram  stainiiij,^  is  shown  by 
the  fact  that  trypsin  will  digest  killed  bacteria  which  are  Gram- 
negative,  but  not  Gram-positive  forms;  gastric  juice  attacks  only  a 
few  Gram-positive  bacteria.-^  They  are  also  more  resistant  to  alka- 
lies, 1  per  cent.  KOH  dissolving  only  the  Gram-negative  bacteria. 
Brundy  -^  considers  that  they  are  more  permeable  to  iodin,  so  that  a 
more  central  iodin-dye  precipitate  occurs,  and  Eisenberg  ^'-^  suggests 
that  lipoid-protein  compounds  in  the  surface  are  important,  in  support 
of  which  is  the  observation  that  ether  extraction  of  staphylococci  ren- 
ders them  negative  to  Gram's  method,  while  colon  bacilli  treated  with 
lecithin  become  positive.^"  Jobling  and  Petersen  -^-^  have  also  found 
the  lipoids  of  Gram-positive  bacteria  more  resistant  to  extraction  by 
fat  solvents  than  lipoids  of  Gram-negative  bacteria,  and  Tamura^*"' 
found  that  the  lipoid  extract  contains  the  bacterial  element  respon- 
sible for  Gram  staining.  The  first-named  authors  suggest  a  relation 
between  the  high  content  in  unsaturated  fatty  acids,  w^ith  their  high 
affinity  for  iodin,  and  the  positive  Gram  staining.  On  the  other  hand, 
llottinger  ^°''  attributes  Gram  staining  solely  to  the  degree  of  disper- 
sion of  the  nucleo-proteins,  which  he  believes  to  be  higher  in  the 
Gram-negative  forms.  Benians  ^^  has  found  that  crushed  Gram-posi- 
tive bacteria  are  promptly  decolorized,  indicating  that  the  dye  and 
the  cell  contents  do  not  form  an  insoluble  compound,  but  that  the  bac- 
terial cell  wall  is  the  chief  factor  in  determining  Gram  positiveness ; 
presumably  the  iodin  renders  the  cell  membrane  impermeable  to  al- 
cohol. This  important  contribution  has  been  confirmed,  as  far  as  the 
staining  of  tubercle  bacilli  is  concerned,  by  Hope  Sherman,^'  who 
corroborates  the  finding  of  Benians  that  if  the  bacilli  are  not  intact 
they  are  neither  acid  fast  nor  Gram  positive.  The  same  is  true  of 
yeast  cells  (Henrici )."-•'' 

\  BACTERIAL  ENZYMES  ^^ 

The  metabolic  processes  of  bacteria  seem  to  be  closely  dependent 

27  Btircrers,  Sehermann  and  Sclireiber,  Zeit.  f.  Hyg.,  1011  (70),  110;  Weinkoff, 
Zeit.  Immunitat..   ] 012    (11).  1. 

28  Cent.  f.  13akt.,  ii  Abt..  1008   (21),  62. 
2!)  Cent.  f.  Bakt..  1010  (56),  10.3. 

30  Jour.  Path,  and  Bact.,   1011    (16).   146. 
30a  Zeit.  phvsiol.  Chem.,  1014    (SO),  280. 
30b  Gent.  f.'Bakt.,  1016   (76),  367. 

31  Jour.  Path,  and  Bact..  1012    (17).  100, 

32  .Tour.  Infec  Bis.,  1013    (12),  240. 
32a  .Tour.  yiod.  Pes..  1014    (30).  400. 

33  See  Fuhrniann  ("Vorlosungen  iiber  Baktcrienenzvme,"  Jena,  1007)  for  com- 
plete bibliography  to  that  date. 

8 


114  CUEMJt^TRY  OF  RACTERIA    A\D   THEIR  PRODUCTS 

upon  enzyme  action,  just  as  with  lii<i:lier  cells.  Liquefaction  of 
jielatiu  is  a  familiar  example  of  the  enzyme  action  of  bacteria;  and 
since  the  filtered  cultures  of  liquefactive  bacteria  are  also  capable 
of  digestintj:  gelatin,  tlie  enzymes  are  evidently  excreted  from  the 
cells.  Dead  baeteria,  killed  by  thymol  or  by  other  antiseptics  that 
do  not  destroy  proteolytic  enzymes,  will  also  digest  gelatin.  Numer- 
ous investigations  have  established  the  wide-spread  occurrence  of 
many  soluble  enzymes  both  in  bacteria  and  in  their  secretions,  indi- 
cating that  bacterial  cells  are  as  dependent  on  enzymes  for  the  pro- 
duction of  their  metabolic  activities  as  are  higher  types  of  cells,  and 
that  these  enzymes  are  not  onlj-  present  as  intracellular  constituents, 
but  that  they  also  escape  from  the  cells.  Even  the  spores  may  con- 
tain active  enzymes.^*  A  striking  property  of  bacteria  is  their  re- 
ducing power,  whicii  has  led  to  the  introduction  of  selenium  and 
tellurium  salts,  which  are  reduced  to  the  metals,  as  an  index  of  bac- 
terial life  and  activity  (Gosio). 

The  diffusion  method  of  Wijsman,  or,  as  it  is  more  frequently 
called,  auxanographic  method  of  Beijerinck,  offers  a  relatively  simple 
)neans  of  detecting  the  presence  of  extracellular  bacterial  enzymes. 
Eijkman^"^  in  particular  has  used  this  method,  which  consists  of  mix- 
ing agar  with  milk,  or  starch,  or  whatever  material  is  to  serve  as  the 
indicator  of  the  enzyme  action  ;  the  agar  is  then  inoculated  with  bac- 
teria and  plated  (or  else  the  bacteria  are  inoculated  as  a  streak  on 
the  surface  of  the  agar).  About  each  colony  there  will  appear  a 
zone  of  clearing  in  the  medium,  if  it  produces  enzymes  digesting 
the  admixed  substance.  By  this  means  Eijkman  found  that  all  bac- 
teria that  produce  enzymes  digesting  gelatin  also  digest  casein,  and 
those  that  do  not  digest  gelatin  are  equally  without  effect  on  casein ; 
therefore,  it  is  probably  the  same  enzyme  that  digests  both.  As  the 
hemolytic  action  of  bacteria  is  not  constantly  related  to  their  gelatin- 
dissolving  i)ropei'ty,  the  hemolysis  probably  is  produced  by  other 
means  than  the  proteolytic  enzymes.''"  A  few  pathogenic  bacteria 
(anthrax,  cholera)  digest  starch,  and  B.  pyocyaneus,  Staphylococcus 
pyogenes  aureus,  and  B.  prodigiosus  all  produce  fat-splitting  enzymes 
demonstrable  by  this  method.  B.  pyocyaneus,  Eijkman  found,  di- 
gested elastic  tissue  readily,'''  as  also  did  a  bat-illus  i-cseinbling  B. 
suhtiiis  obtained  from  the  tissue  of  a  gangreiious  lung.  The  following 
table  by  Buxton ''^  gives  an  idea  of  the  disti-ihution  of  enzymes  in 
bacterial  secretions  as  determined  by  the  auxanographic  method : 

•'<4  EfTront,  Mon.  sc.  Quesnevillc   I!i(i7,  j).  Sl. 

35  Cent.  f.  Biikt.,  1001    (20),  S4  1 . 

3«  Sec  .Jordan.   Biol.  Stiulicy  1)\    iIil'  |.u|>i!s  of  W.  T.  Sod^^wick.   r.KMI.  p.    IJ). 

37  (Vnl.  f.   Hakl..   1 !»(»:!    (:(.-,).  'l . 

3«  Aiiicricaii  .Med.,  l!l(i:{    ( (l ) .  i:i7. 


BACTERIAL  ES'/A  MKS 


115 


ENZYMES   IIYDKATIXG   CARBOHYDRATES  39 

Amylase.  Maltase.  Invertase.  Lactase. 

1.  Antlirax + 

2.  Cliolera + 

'^.  Coli   foiniiiunis — 

4.  'ryi)lu)i(l — 

5.  l)i|)litlioria — 

(i.  Slapli.   ])yo<;eiios   aureus  — 

7.  J.ac-tis    aerogenes       ....      — 

8.  Pyocyaiieus — 

0.  Violacoii.s — 

1(K   Mycoidcs  + 

11.  Prodigiosus — 

12.  Saecharomyces  niger      ...  — 

13.  Saecharomyces    neoformaus  — 

14.  Aspergillus    niger     .      .      .      .  + 

15.  Aspergillus  oryzoe    .      .      .      .  + 

PROTEOLYTIC   ENZYMES,   DIGESTING 


Inulase. 


+ 
+ 
+ 
+ 

+ 
+ 


+ 


+ 


+ 


+ 


+ 

+ 

+ 

+ 

+ 

+ 

— 

+ 

Milk. 

Gela- 
tin. 

Serum 

Egg- 
albu- 
men. 

Fibrin. 

Red 
blood- 

Coagul. 

Diges- 
tion. 

corpus- 
cles. 

+ 
+ 

+ 

+ 

+ 

+ 

+ 
+ 

+ 

+ 

+ 

+ 

+ 

+ 
+ 

+ 

+ 
+ 
+ 

+ 
+ 
+ 

+ 

+ 

+ 
+ 

+ 

+ 

+ 
+ 

+ 

+ 
+ 

+ 

-4- 

2.   Cholera      

+ 

3.  Coli   communis    

4.  Staph,    pyogenes    aureus. 

5.  Streptococcus     pyogenes. 

6.  Pyocyaneus      

+ 
+ 

+ 

8     Mvcoides      

+ 

+ 

10.  Aspergillus  niger    

11.  Aspergillus    oryzoe     .... 

+ 

Rennin  is  produced  by  many  bacteria,  as  is  shown  by  their  coagu- 
lating milk,  independent  of  any  acid  reaction,"  and  protease  from 
pyocyaneus  causes  "pilastein"  formation  in  albumose  solutions  (Zak).*^ 
Bacteria  which  give  negative  results  by  the  plate  method  may  con- 
tain active  lipase  demonstrable  in  killed  bacteria  by  direct  action 
upon  fats  and  esters,  these  lipases  behaving  exactly  like  the  lipase  of 
animal  tissues  (Wells  and  Corper)  ;  ■*-  staphylococcus  and  pyocyaneus 
are  more  actively  lipolytic  than  B.  coli,  B.  dysenteri<t  and  B.  tuber- 
culosis. 

Schmailowitsch  *^  stated  that  the  amount  and  nature  of  enzymes 
produced  hy  bacteria  is  modified  by  the  amount  and  nature  of  their 
food,  but  Jordan  found  that  gelatinase  is  produced  by  bacteria  grow- 
ing on  non-protein  media;  he  failed  entirely  to  support  the  statement 
of  Abbott  and  Gildersleeve  **  that  bacteria  grown  on  gelatin  produce 
)nuch  more  active  gelatin-dissolving  enzyme  than  do  bacteria  grown 

39  III  relation  to  carbohydrate  enzymes,  the  extensive  studies  of  Kendall  (Jour. 
Biol,  ('hem.,  1912,  vol.  12)  should  be  con.sulted.  He  empliasizes  especially  tiiat  as 
a  rule  bacteria  ferment  carboliydrates  in  preference  to  attacking  proteins  wlien 
botli  foodstutt's  are  availabh>. 

40  Contradicted  bv  DeWaele,  Cent.  f.  Bakt.,  1905    (39),  353. 

41  Hofmeister's  Beitr.,   1907    (10),  287. 

42  Jour.  Infect.  Dis.,  1912  (11),  388;  literature  on  bacterial  lipases.  See  also 
Kendall. 

43  Wratschebnaja  Gazetta,  1902.  p.  52. 

44  Jour.  Med.  Research,   1903    (10),  42. 


]16  CHEMISTRY  OF  BACTERIA  AND  THEIR  PRODUCTS 

on  bouillon.  There  does  not  seem  to  be  any  important  relation  be- 
tween enzj'me  production  and  pathogenicity.*^" 

In  general,  bacterial  proteolytic  enzymes  resemble  trypsin  more 
closely  than  they  do  pepsin,  acting  best  in  an  alkaline  medium;  but 
the  enzymes  extracted  from  bacterial  cultures  are  very  feeble  as  com- 
pared with  pancreatic  trypsin.  It  is  probable  that  there  are  several 
distinct  proteolytic  enzymes  in  bacterial  cells,  gelatinase  being  a  dis- 
tinct protease  (Jordan)."''  Abbott  and  Gildersleeve  found  that  the 
gelatin-dissolving  enzyme  of  bacteria  resists  a  temperature  of  100°  C. 
for  as  long  as  fifteen  to  thirty  minutes,  but  Jordan  found  that  the 
reaction  of  the  medium  modified  greatly  this  heat  resistance. 
Schmailowitsch  '^^  states  that  some  bacteria  produce  an  enzyme  acting 
in  acid  medium  upon  gelatin  but  not  upon  albumin,  and  this  enzyme 
carries  the  digestion  only  as  far  as  the  gelatin-peptone  stage,  whereas 
the  enzymes  acting  in  an  alkaline  medium  carry  the  splitting  through 
to  leucine,  tyrosine,  etc.  Kendall  and  Walker  ^^-'^  state  that  the  pro- 
teolytic enzymes  of  B.  pruteus  are  not  formed  when  the  bacteria  have 
enough  carbohydrate  supplied  so  that  they  need  not  depend  on  pro- 
teins for  their  energy  requirements;  deaminization  is  independent  of 
proteolysis  and  represents  intracellular  enzj^me  action.  Plenge  *" 
suggests  that  there  is  a  special  enzyme  digesting  nucleoproteins. 
Bacteria  are  able  to  split  nucleic  acids  and  to  convert  amino-purines 
into  oxypurines,  but  they  do  not  carry  the  oxidation  to  uric  acid; 
putrefactive  bacteria  can  slowly  destroy  uric  acid  (Schittenhelm),*^ 
and  B.  coli  destroys  purines.*'^^ 

Cacace  *^  investigated  the  splitting  products  of  gelatin  and  coagu- 
lated blood  when  digested  by  B.  anthracis,  Staph,  pyogenes  aureus, 
and  Sarcina  aurantiaca,  and  found  that  proteoses  and  peptone  are 
produced,  which  disappear  in  the  later  stages  of  digestion.  Rettger  *^ 
found  leucine,  tyrosine,  tryptophane,  as  well  as  phenols,  skatole,  in- 
dole, aromatic  oxy-acids,  and  mercaptan,  among  the  products  of  bac- 
terial decomposition  of  egg-albumen  and  meat;  proteoses  and  pep- 
tones appear  in  the  early  stages,  but  later  disappear,  as  also  eventu- 
ally do  the  leucine,  tyrosine,  etc.  Choline  has  also  been  found  in  the 
l)roducts  of  autolysis.^" 

The  digestive  power  of  the  filtrates  of  cultures  and  of  killed  bac- 
teria is  far  less  than  that  of  the  living  bacteria  (Knapp).^^     Strepto- 

4«a  Rosenthal  and  Patai.  Cent.  f.  Bakt.,  1014   (7.'?),  400;    (74),  3011. 
4-Ji)  Corroborated  bv  Bertiau,  Cent.  f.  Bakt..  1914   (74),  374. 
4.-.  Abst.   in   Biocliein.  Centr.,   1903    (1),  230;    see  also  DeWaele.  Cent.   f.  Bakt., 
1905    (39),  .353. 

45a.l,Hir.  Infect.  Dis..  1915    (17),  442. 
40Zeit.  f.  i.livsiol.  C'liem.,   1903    (39),   190. 

47  Zeit.  physiol.  Ciiem.,  190S    (57).  21. 

4Ta  Siven,  Zeit.  phvsiol.  Chem.,  1914    (91),  33(). 

48  Cent.  f.  Bakt..  1901    (30),  244. 
40Amer.  Jour,  of  Physiol.,  1903    (8),  284. 

r.o  KiitHcher  and  Loliiiiann,  Zeit.  plivsiol.  Ch(>ni.,  1903    (39),  313. 
M  Zeit.  f.  lleilk.    (Cliir.  Abt.)    1902   (23),  230. 


IMMU^aiTY  AGAiysr  BACTEIilAh  ES/AMEfi  117 

cocci  digest  proteins  of  exudates  feebly,  staphylococci  more  rapidly, 
and  colon  bacilli  are  still  more  active.  He  could  find  no  relation 
between  the  proteolytic  power  of  the  bacteria  and  the  severity  of  the 
infection  from  which  they  came.  Staphylococci  can  cause  coagula- 
tion of  pla.sma  and  then  dissolve  the  coagulum,  showing  the  presence 
of  two  enzymes,  staphylokiimse  and  fihrinolysin  (Kleinschmidt).^^ 
Sperry  and  Rettger,"*  however,  found  that  even  the  most  actively 
putrefactive  bacteria  are  unable  to  attack  or  grow  upon  carefully 
purified  proteins,  although  the  presence  of  small  amounts  of  peptone 
or  other  available  nutrient  makes  the  proteins  available  to  the  bac- 
teria; apparently  they  must  have  some  nutrient  more  available  than 
intact  protein  molecules  to  enable  them  to  grow  sufficiently  to  produce 
enough  free  enzymes  to  attack  the  proteins.  By  virtue  of  their  pro- 
teolytic enzymes,  filtrates  of  bacteria  that  liquefy  gelatin  also  can 
digest  hardened  liver,  kidney  and  other  tissue  elements  in  vitro,  the 
changes  resembling  those  of  necrobiosis.^-^ 

Catalase  is  demonstrable  in  bacteria,  the  anaerobic  forms  showing 
the  least  activity  (Rywosch),^^  but  practically  no  species  is  entirely 
inactive  ( Jorns)  ;  ^*  it  may  exist  as  either  endo-  or  ecto-enzyme.  Cer- 
tain bacteria  and  actinomyces  exliibit  oxidative  effects,  resembling 
tyrosinase,  but  such  an  enzyme  could  not  be  extracted  by  Lehmann 
and  Sano.^^ 

Immunity  against  bacterial  enzymes  may  be  secured  as  it  is  against 
other  enzymes.  Abbott  and  Gildersleeve  ^*  found  that  by  injections 
into  animals  of  proteolytic  bacterial  filtrates  which  were  only  slightly 
toxic,  the  serum  of  the  animals  acquired  a  slight  but  specific  in- 
crease in  resistance  to  the  proteolytic  enzymes  of  the  filtrates.^^ 
Normal  serum  contains  a  certain  amount  of  enzjTne-resisting  sub- 
stance. Other  observers  have  found  that  immunization  against  living 
or  dead  bacteria  leads  to  the  production  of  substances  antagonistic 
to  their  enzj-mes,  but  the  degree  of  resistance  acquired  is  never 
great,  v.  Dungern  ^'  found  that  the  serum  of  animals  infected  with 
various  bacteria  prevented  digestion  of  gelatin  by  the  enzymes  ob- 
tained from  cultures  of  the  same  species  of  bacteria.  He  applied 
this  fact  to  the  diagnosis  of  infectious  conditions,  finding  that  the 
serum  of  a  patient  with  osteomyelitis  was  over  twenty  times  as 
strongly  inhibitory  to  staphylococcus  enzymes  as  was  serum  of  nor- 
mal persons.  The  reaction  is  specific,  cholera  vibrio  enzjTiies  not  be- 
ing inhibited  to  any  corresponding  degree. 

52Zeit.  Immunitat.,  1900    (3).  516. 
52a  .Jour.  Biol.  Chem.,  1915    (20),  445. 
52bBittrolff.  Ziegler's  Bcitr.,  1915   (60),  3.37. 

53  Cent.  f.  Bakt.,  1907    (44),  295. 

54  Arch.  f.  Hvcr.,  1908   (67),  134. 

55  Arch.  f.  Hyg.,  1908    (67),  99. 

56  Antigelatinase  has  also  been  obtained  bv  Bertiau.  Cent.  f.  Bakt.,  1914  (74), 
374. 

57  Munch,  med.  Woch.,  1898    (45),  1040. 


118  CHEMISTRY  OF  BACTERIA   A\D  THEIR  PRODUCTS 

Kaiitorowicz  ""''  and  de  AVaele  ''•'  state  tliat  bacteria  contain  an  in- 
tracellular anti-protease  wiiicli,  with  most  bacteria,  holds  in  check 
the  proteolytic  action;  only  with  tlie  liquefying  bacteria  are  the 
proteases  in  excess.  Bacteria  grow  well  in  strong  solutions  of  en- 
zymes, and  without  destroying  the  enzymes  (Fermi).®"  After  Gram- 
negative  bacteria  have  been  heatetl  to  80°  they  are  readily  digested  by 
trypsin,  pepsin  or  leucocytic  proteases;  but  Gram-positive  bacteria 
are  resistant  even  after  heating.  This  is  ascribed  by  Jobling  and 
Petersen""''  to  the  unsaturated  fatty  acids,  wliicli  are  present  in 
greater  amounts  in  Gram-positive  bacteria. 

Autolysis  of  Bacteria. — Autolysis  occurs  also  in  bacteria,  their 
proteolytic  enzymes  digesting  the  cell  substance  whenever  the  organ- 
isms are  killed  by  agents  (chloroform,  toluene,  etc.)  that  do  not  de- 
stroy these  enzymes,  and  which,  being  fat  solvents,  may  facilitate  di- 
gestion by  removing  the  inhibitory  lipoids.  Even  the  absence  of  food 
leads  to  autolysis,  presumably  because  the  normally  existing  auto- 
lytic  processes  are  not  counteracted  by  synthesis  of  new  protein  ma- 
terial ;  hence,  autolysis  occurs  M^hen  bacteria  are  placed  in  salt  solution 
or  distilled  water.  Although  it  had  been  known  for  many  years  that 
yeast  cells  digest  one  another  when  there  is  nothing  else  for  them  to 
live  upon,  the  first  definite  study  of  bacterial  autolysis  seems  to  have 
been  made  by  Levy  and  PfersdorfP  "^  and  Conradi.^-  The  former 
digested  anthrax  bacilli  (in  whose  bodies  are  contained  rennin,  lipase 
and  protease)  under  toluene  for  several  weeks  and  obtained  a  slightly 
toxic  product.  Conradi  permitted  dysentery  bacilli  and  typhoid 
bacilli  to  digest  themselves  in  normal  salt  solution  for  twenty-four  to 
forty-eight  hours  at  37°  C,  and  obtained  in  this  way  the  soluble, 
highly  poisonous  endotoxins  of  the  bacteria,  which  are  liberated  by 
the  destruction  of  the  bacterial  structure  by  the  autolytic  enzymes. 
Longer  autolysis  results  in  the  destruction  by  the  enzymes  of  the  en- 
dotoxins themselves.  Rettger  '''^  found  among  the  autolytic  products 
of  bacteria,  leucine,  tyrosine,  basic  substances,  and  phosphoric  acid. 
Under  favorable  conditions  complete  autolysis  can  occur  in  two  to 
ten  days. 

Brieger  and  IMayer  "*  found  that  at  room  temperature  (15°  C.) 
practically  no  autolysis  occurs  with  typhoid  bacilli  in  distilled  water, 
and  the  soluble  products  thus  obtained  are  quite  non-toxic,  although 
if  injected  into  animals  they  give  rise  to  the  production  of  agglu- 
tinins   and    bacteriolysins.     Bertarelli  '''■'    has    used    tlie    products    of 

ssMiinfh.  mod.  Woch.,  1909   (.'56),  897. 
•'•oCont.  f.  Bakt.,  1909   (50),  40. 
00  Arch.  Farmac'ol.,   1909    (S),  481. 
«oa,Tour.  Kxy).  Med.,   1914    (20),  321. 
«i  Dc'ut.  nicd.   Wocli..   1902    (2S),   879. 
12  Ibid.,   ]'M):\    (29),  2(). 
03  Jour.  :M('d.   Kcsoaroli,    1904    (13),   79. 
«4Dcut.  iiK'd.  WOcli.,  1904    (.30),  !)S0 
05  Cent.  f.   Bakt.,    1905    (38),  5S4. 


.\(  Toi.Ysis  OF  ii.\("n:ni.\  110 

autolysis  of  cliolcra  vibrios  sncccssi'uily  in  the  j)i'0(hicti()i)  of  iiniiiu- 
iiity,  and  states  tliat  the  i)ro(lu('ts  of  autolysis  consist  larfi;('ly  of  nu- 
c'leins. 

It  is  ])i'ol)al)l('  tliat  in  cvciy  culture  jjactcria  arc  couslantly  Ijcinfj 
destroyed,  eitlier  by  their  own  enzymes  or  by  tlie  proteolytic  enzymes 
of  the  other  bacteria.  Some  bacteria  are  much  more  rapidly  auto- 
lyzed  tliau  others,  cholera  vibrios,  colon,  typhoid,  and  dysentery 
bacilli  being  rapidly  di<iested,  while  sti'cptococci,  staphylococci  and 
tubercle  bacilli  are  very  little  and  slowly  autolyzed.  In  p:emn-al,  the 
Gram-positive  organisms  resist  autolysis  longest. 

Conradi,'^*'  who  has  shown  that  certain  products  of  autolysis  of  tis- 
sues are  bactericidal,  believes  that  also  in  cultures  powerfully  bacteri- 
cidal substances  are  produced  through  autolysis  of  the  bacteria.  This, 
he  tliinks,  accounts  for  the  decrease  in  numbers  of  living  bacteria  that 
always  sets  in  after  a  short  period  of  growth  on  artificial  media;  but 
there  is  much  doubt  as  to  these  substances  being  of  any  considerable 
importance  in  the  body."^  It  has  been  found  by  Turro  '''^  that  ex- 
tracts from  various  tissues  containing  autolytic  enzymes  can  digest 
bacterial  cells.**^  It  is  very  possible  that  the  endotoxins  contained 
Avithin  such  pathogenic  bacteria  as  typhoid  and  cholera  are  liberated 
through  digestion  of  the  bacteria,  either  by  autolysis  or  b.v  the  en- 
zymes of  the  leucocytes  and  tissues  of  the  organism  that  they  have 
infected.  These,  and  a  number  of  other  bacteria,  produce  no  soluble 
toxins  that  dififuse  from  the  cells  as  do  diphtheria  and  tetanus  toxin, 
and  it  is  difficult  to  explain  the  toxic  effects  these  bacteria  produce 
without  assuming  that  their  intracellular  toxins  are  liberated  in  some 
such  way.  It  is  also  quite  probable  that  the  enzymes  found  in  fil- 
trates from  bacterial  cultures  are  liberated  from  the  bacterial  cells 
only  when  these  have  been  autolyzed. '■°  "With  the  possible  exception . 
just  mentioned,  there  is  little  evidence  that  the  bacterial  enzymes 
play  any  important  role  in  infectious  diseases.  They  may  be  a  slight 
factor  in  the  digestion  of  tissue  and  exudates  in  suppuration,  but  as 
compared  with  the  leucocytic  enzymes  their  influence  is  probably  mi- 
nute; beyond  this  they  have  no  apparent  influence  upon  their  host, 
and  are  chiefly  concerned  in  the  metabolism  of  the  bacteria.  The 
proteoses   and   peptones   produced   by   bacterial    action    and    isolated 

«oMiineh.  med.  Wochenschr.,   1905    (r)2),   1761. 

67  See  Eiikman,  Cent.  f.  Bakt.,  1906  (41),  31)7:  Passini,  Wieii.  klin.  Wopli., 
1906    (19),  627. 

08  Cent.  f.  Bakt.,  1902    (.32),  105. 

'!«  Sigwart  ( Arl).  a.  d.  Path.  Inst.  Tiibingen,  1902  (3),  277)  found  tliat  trypsin 
and  pepsin    (without  acid)   do  not  injure  living  anthrax  baeilli. 

70  Emmerich  and  Lrew  (Zeitsclir. '  f.  ITyg.,  1899  (31),  1),  having  found  that 
pyoci/anase  is  capable  of  destroying  and  digesting  otiier  bacteria  tlian  pyocy- 
aneus,  suggested  that  it  might  be  a  potent  factor  in  producing  artificial  immu- 
nity. Their  rather  remarkable  liypotheses  I'ave  been  much  contested,  and  are 
of  questionable  value.  (See  Petr'ie,  .Jour,  of  Pathol,  and  Bacteriol.,  1903  (8), 
200;  also,  Rettger  (Jour.  Infectious  Diseases,  1905  (2),  562);  Emmerich 
Miinch.  med.  Woch.,   1907    (54),  2217). 


120  CHEMISTRY  OF  BACTERIA  AND  THEIR  PRODUCTS 

from  cultures  do  not  seem  to  be  any  more  toxic  than  those  produced 
by  pepsin  and  trypsin,  but  violent  poisons  may  be  liberated  from 
bacteria  during-  autolj'sis,  as  Rosenow  '^  has  shown  for  the  pneumo- 
coccus  and  other  bacteria;  these  poisons  seem  similar  to  or  identical 
with  the  so-called  anaphylatoxin  which  is  supposedly  formed  by  the 
difrestion  of  bacteria  witli  serum  complement,  and  presumably  they 
are  proteoses  or  polypeptids,  but  their  exact  nature  is  not  known. 
(See  Anaphylatoxin,  Chap,  vii.) 

POISONOUS  BACTERIAL  PRODUCTS 

Almost  without  exception  all  the  harm  that  bacteria  do  is  brought 
about  by  means  of  the  chemical  substances  produced  in  one  way  or 
another  by  their  metabolic  processes.  Animal  parasites  may  do  hainn 
mechanically,  but  with  the  possible  exception  of  the  efifects  of  capil- 
lary emboli  (especially  with  anthrax),  bacteria  produce  all  their  ef- 
fects through  chemical  means.  The  poisonous  chemical  substances 
produced  by  bacteria  are  commonly  grouped  into  four  classes : 

I.  Products  of  the  decomposition  of  the  media  upon  which  the  bac- 
teria are  growing;  among  these  the  best  known  are  the  ptomdins. 

II.  Soluble  poisons  manufactured  by  the  bacteria,  and  secreted 
from  the  cell  into  its  surrounding  media — the  true  toxins. 

III.  Poisons  manufactured  by  the  bacteria  which  do  not  escape 
from  the  normal  cell  but  which  are  as  specific  in  their  poisonous  prop- 
erties as  the  true  toxins;  because  of  their  intracellular  situation  they 
are  called  endotoxins. 

IV.  Poisonous  protein  constituents  of  the  bacterial  cell,  which  form 
part  of  the  cell  protoplasm,  but  which  are  not  soluble,  and  the  poison- 

■  ous  effects  of  which  are  not  specific  and  not  usually  responsible  for 
the  disease;  these  are  called  hactcrial  proteins. 

PTOMAINS 
Ptomai'ns,  the  soluble  basic  nitrogenous  substances  that  are  found 
in  the  medium  in  which  bacteria  have  been  growing,  were  the  first 
bacterial  products  that  were  recognized,  and  for  some  time  it  was 
believed  that  it  was  through  the  production  of  such  alkaloid-like  sub- 
stances that  bacteria  caused  disease,  just  as  poisonous  plants  owe 
their  effects  to  poisonous  alkaloids.  It  was  soon  found,  liowever,  tliat 
the  ptomains  that  could  be  isolated  from  cultures  of  pathogenic  bac- 
teria were  insufficient  by  their.selves  to  cause  the  poisonous  effects 
that  such  cultures  produced  when  injected  into  animals.  The  isolated 
ptomains  were  not  only  far  less  poisonous  than  the  original  culture, 
but  furthermore  they  did  not  produce  the  .symptoms  and  anatomical 
changes  characteristic  of  the  diseases  that  the  pathogenic  organism 

71  .Tour.  Infect.  Dis.,  1912    (10),  113;    (11),  94.  235  and  480. 


PTOMAJXS  121 

caused.  Moreover,  the  majority  of  ptomai'ns  are  not  very  poisonous, 
and  highly  poisonous  ptomains  may  be  produced  by  non-pathogenic 
bacteria.  As  a  result,  the  work  on  ptomains,  which  once  occupied 
man}'  laboratories  and  promised  to  reveal  tlie  entire  chemistry  of  bac- 
terial intoxication,  has  now  been  almost  completely  dropped.  The 
interest  in  ptomains  is  by  no  means  entirely  historical,  however,  for 
poisonous  ptoinai'iis  at  times  do  enter  the  body  and  cause  illness, 
sometimes  even  death.  The  close  chemical  resemblance  to  vegetable 
alkaloids  of  some  of  the  ptomains  that  may  arise  in  decomposing 
corpses,  makes  them  of  great  importance  to  chemists  searching  for 
the  cause  of  death  in  cases  of  supposed  poisoning.  Tlierefore  the 
most  essential  features  of  the  ptomains  and  their  chief  known  rela- 
tions to  intoxications  will  be  briefly  discussed,  refeiTing  the  reader 
for  a  full  consideration  to  Vaughan  and  Novy's  "Cellular  Toxins" 
and  Barger's  "The  Simpler  Natural  Bases." 

The  ptomains  owe  their  basic  character  to  nitrogen-containing 
radicals,  principally  amino-groups,  and  hence  are  formed  from  ni- 
trogenous substances,  chiefly  proteins,  which  contain  their  nitrogen 
in  the  amino  form.  Probably  most  ptomains  arise  from  the  decompo- 
sition of  the  protein  medium  upon  which  the  bacteria  grow,  although 
undoubtedly  part  of  the  ptomains  is  also  formed  from  the  destruc- 
tion of  the  bacterial  cells  themselves;  how  large  a  part  of  the  pto- 
mains is  formed  by  intracellular  bacterial  processes  and  how  much 
by  cleavage  of  the  proteins  of  the  media  by  extracellular  bacterial 
enzymes  is  unknown.  The  structure  of  the  ptomains  shows  them  to 
be  very  closely  related  to  the  amino-aeids  obtained  by  cleavage  of  the 
protein  molecule  by  enzymes  and  other  hydrolytic  agencies ;  and  the 
determination  of  the  composition  of  the  several  amino-aeids  of  the 
proteins  has  quite  cleared  up  the  problem  of  the  origin  of  the  pto- 
mains. Presumably  these  secondary  changes  result  from  the  action 
of  special  enzymes  upon  the  amino-aeids.  ]\Iost  of  the  ptomains  are 
free  from  or  poor  in  oxygen,  hence  reduction  processes  are  probably  im- 
portant in  their  production.  The  poisonous  ptomains.  which  are  de- 
cidedly in  the  minority  among  the  entire  group,  are  themselves  subject 
to  decomposition,  being  most  abundant  in  the  cultures  after  a  certain 
period  of  time,  and  then  decreasing  in  amount.  Very  old  cultures  show 
almost  none  of  the  higher  molecular  forms  of  nitrogen,  such  as  pto- 
mains, these  substances  having  been  changed  into  ammonium  and  ni- 
trate compounds.  In  sharp  contradistinction  to  the  toxins,  the  pto- 
mains are  hy  no  means  specific.  No  matter  upon  what  medium  diph- 
theria bacilli  grow,  the  toxin  produced  has  qualitatively  the  same  prop- 
erties, whereas  the  nature  of  the  ptomains  depends  not  only  upon  the 
nature  of  the  bacteria  producing  them,  but  also  even  more  upon  the  sort 
of  soil  upon  which  the  bacteria  are  grown,  the  temperature,  the  dura- 
tion of  the  process,  and  the  quantity  of  oxygen  furnished.     The  same 


122  CHEMISTRY  OF  BACTERIA  AXD  THEIR  PRODUCTS 

orgranism  may  produce  totally  different  ptoinauis  when  grown  on 
different  media  or  under  different  conditions.  Another  essential  dif- 
ference is  that  w^e  cannot  obtain  an  immune  serum,  antagonizing  the 
action  of  ptomains,  by  injecting  ptomai'ns  into  animals. 

Ptomains  are  chiefly  the  cause  of  disease  when  they  are  taken 
in  with  food  in  which  thej'  have  been  produced  by  bacterial  decom- 
position. Besides  this  food  poisoning,  it  is  also  possible  that  pto- 
mains may  be  formed  by  putrefaction  within  the  gastrointestinal 
tract.  Another  possible  source  of  ptomains  is  furnished  by  decom- 
posing tissues  in  gangrene.  It  is  doubtful  if  ptomains  are  produced 
in  sufficient  quantities  by  pathogenic  bacteria  infecting  living  tissue 
to  be  of  any  importance.  Food  poisoning  is  by  no  means  uncommon, 
but  it  is  not  always  due  to  ptomains ;  it  may  be  the  result  of  poisonous 
materials  contained  abnormally  in  the  food,  that  are  not  ptomains, 
e.  g.,  botulism ;  or  it  may  be  due  to  an  infection  of  the  animal  from 
which  the  meat  came  with  pathogenic  organisms,  particularly  the 
B.  enteritidis  of  Gaertner  and  other  bacteria  related  to  the  colon- 
typhoid  group ;  or  in  other  ways  food  ordinarily  wholesome  may  be- 
come poisonous.  The  commonest  sources  of  ptomai'n  poisoning  are 
imperfectly  preserved  canned  meats,  sausages,  decomposing  fish, 
cheese,  ice-cream,  and  milk.'^ 

Chemical  Composition  of  Ptomains. — To  indicate  the  composi- 
tion and  nature  of  i)tomains  a  few  of  the  more  important  ones  will  be 
described.     As  illustrative  of  the  simpler  forms  may  be  mentioned: 

Methyl  amine,  CH3— JSTH,. 

Dimethyl    amine,  CH3— NH— CH3. 

Tri-methyl    amine,  CH3— N— CH3. 

CH3. 

These  bodies,  which  are  commonly  found  in  decomposing  proteins, 
are  but  very  slightly  toxic,  and  of  little  pathological  importance. 

The  source  of  the  ptomains  in  the  various  amino-acids  is  usually 
easily  traced  through  their  chemical  structure,  and  Ackermann  and 
Kutscher  '*  have  classified  them  in  this  relation  under  the  name 
"  aporrhegma." 

When  we  examine  the  structural  formula^  of  some  of  the  larger 
l)tomain  molecules  and  compare  them  with  the  formula'  of  the  amino- 
acids  that  form  the  protein  molecule,  the  relation  is  apparent,  e.  g., 
compare  iso-amylamine  with  leucine. 

CH3.  CH3 

„„  >  CH— CH,— CH,— NHj  ^„  >CH— CH,— CH— NH, 

CHa^^  CJI3/  X 

(iso-amylamine)  (leiicinp)       ^('OOH. 

73  All  tlipse  matters  are  discussed  at  Icny^th  hy  ^  an^'liaii  and  Xovy.  to  wliose 
hook  llie  reader   is  referred. 

-•Zeit.  physiol.  riiem.,   1910   (69),  205. 


CIIOLIM-:  a  ROUP  123 

Putrescine,  C^HjoN^,,  structural  formula, 

NH,— CH„— CH:;— CH„— CH,— XI  r,, 

and  cadaverine,  C-II14N.,,  structural  formula, 

NHo— CHo— CH,— CH,— CH,— CH,— NH„ 

are  of  interest  because  they  have  been  found  in  the  intestinal  con- 
tents, arising"  from  putrefaction  of  proteins,  and  also  are  sometimes 
present  in  the  urine  in  cystinuria.'''  They  are  closely  related  to  the 
diamino-acids,  lysine  and  ornithine.  They  are  but  slightly  toxic, 
although  capable  of  causing  local  necrosis  when  injected  subcutane- 
ously.  (See  further  discussion  on  these  and  the  Pressor  Bases  in 
Chap,  xix.) 

The  Choline  Group. — Another  group  of  ptomai'ns,  including  choline 
and  closely  related  substances,  is  also  of  interest.  These  ptomai'ns 
are: 

Choi  ine,  CH^OH— CH„— X  ( CH3 )  3— OH 

Xeurine,  CH„=CH— N  ( CH, )  3— OH 

IMuscarine,  CH  ( OH ) ,— CH„— X  ( CH, )  3— OH 

Betaine,  COOH— CH,— X^  ( CH3 )  3— OH 

The  first  point  of  importance  is  that  choline  is  present  in  every 
cell  normally,  forming  the  nitrogenous  portion  of  the  lecithin  mole- 
cule. Its  source  in  putrefaction  of  tissues  is,  therefore,  plain.  It  is 
possible' that  choline  is  liberated  from  nerve  tissues  when  they  break 
down  in  the  body  during  life,'^®  and  there  is  a  considerable  literature 
on  the  supposed  finding  of  choline  in  the  blood  and  cerebrospinal 
fluid  in  diseases  of  the  central  nervous  system  and  experimental 
lesions  in  nervous  tissues.  At  present  it  seems  probable  that  these 
observations  depend  upon  faulty  methods  of  analysis,  and  it  is  ex- 
tremely doubtful  if  enough  choline  is  ever  set  free  at  one  time  from 
even  severe  acute  nervous  lesions  to  be  detected  in  the  body  fluids  by 
chemical  means.'^"  Hunt  ''-''  has  devised  a  physiological  test  that  per- 
mits of  the  detection  of  as  little  as  0.00001  mg.,  but  he  was  unable  to 
obtain  evidence  that  choline  is  of  any  significance  in  either  physiologi- 
i^al  or  pathological  processes.  Normally  the  largest  amounts  by  far  are 
obtained  from  the  adrenals,  which  also  seem  to  contain  choline  deriva- 
tives of  much  greater  physiological  activity.     Choline  itself  is  some- 

73Udranszkv  and  Baumann,  Zeit.  phvsiol.  Chem.,  ISSn  (1.3),  562:  1880  (15), 
77. 

70  Coriat  (Amor.  Jour,  of  Physiol.,  1904  (12).  .35.3)  has  studied  the  conditions 
\inder  wliich  choline  niav  lie  produced  from  lecitliin.  Putrefaction  of  lecitliin 
or  lecithin-rich  tissues  liberates  choline,  as  also  does  autolysis  of  brain  tissue; 
neither  pepsin  nor  trypsin,  however,  splits  it  from  the  lecithin.  Tn  brain  tissue, 
therefore,  there  seems  to  be  an  enzyme  different  from  trypsin,  whicli  splits 
choline  out  of  the  lecitliin  molecule. 

77  See  Webster,  Biocliem.  .Tour.,  inon  (4),  123;  Kajiura,  Ouart.  Jour.  Exper. 
Physiol.,  inOS  (1),  201;  Handelsmann,  Dent.  Zeit.  Xervenheilk.,  1908  (35),  428; 
Doree  and  Golla,  Biochem.  .Jour.,  1910   (5),  306. 

77a  Jour.  Pharmacol.,  1915   (7),  301. 


124  CHEMISTRY  OF  BACTERIA  AND  THEIR  PRODUCTS 

what  toxic,  but  the  closely  related  body,  neurine,  into  which  it  may  be 
transformed,  is  highl}'  poisonous,  which  makes  choline  an  important 
indirect  source  of  intoxication.  It  is  possible,  for  example,  that 
lecithin  taken  in  the  food  splits  off  choline  in  the  gastro-intestinal 
tract,  and  this  being  converted  into  neurine  gives  rise  to  intoxication 
which  may  be  ascribed  to  food  intoxication.  Likewise  it  has  been 
suggested  that  the  intoxication  of  fatigue  may  be  due,  at  least  in  part, 
to  choline  and  neurine  produced  from  lecithin  decomposed  during  the 
period  of  cellular  activit}^  The  close  structural  relation  to  choline 
and  neurine,  of  the  mushroom  poison,  muscarine,  which  produces 
physiological  effects  very  similar  to  those  of  neurine,  indicates  the 
close  relationship  of  the  putrefactive  ptomains  and  the  vegetable 
alkaloids.  Indeed  a  muscarine  apparently-  identical  with  that  of  the 
mushroom  has  been  found  in  decomposing  flesh,  and  neurine,  presum- 
ably derived  from  lecithin,  may  be  found  in  human  urine. '^^ 
Betaine,  the  fourth  member  of  the  group,  which  has  but  slight  tox- 
icity, is  particularly  well  known  as  a  constituent  of  plant  tissues; 
possibly  betaine  or  other  basic  bodies  may  occur  substituted  for 
choline  in  certain  varieties  of  lecithin  (Lippmann). 

Both  neurine  and  muscarine  are  extremely  poisonous  and  quite 
similar  in  their  effects.  Subcutaneous  injection  of  but  1  to  3  mg. 
of  muscarine  in  man  produces  salivation,  rapid  pulse,  reddening  of 
the  face,  weakness,  depression,  profuse  sweating,  vomiting,  and  di- 
arrhoea. Neurine,  likewise,  causes  salivation,  lachrymation,  vomiting, 
and  diarrhoea.  In  fatal  poisoning  respiration  ceases  before  the  heart 
stops.  Both  poisons  resemble  phj'sostigmine  in  their  stimulation  of 
secretion  and  are  equally  well  counteracted  by  atropine.  The  toxicity 
of  these  substances  is  so  great  that  not  a  large  amount  would  need 
to  be  formed  by  oxidation  of  choline  to  produce  severe  symptoms, 
altliough  it  is  not  kno\\ai  that  this  actually  occurs  in  the  body.  When 
introduced  by  mouth,  the  lethal  dose  of  neurine  is  ten  times  as  great 
as  when  injected  subcutaneously,  indicating  that  chemical  changes  in 
the  gastro-intestiiial  tract  or  liver  offer  some  protection  against  in- 
toxication by  these  substances  when  taken  in  tainted  food.  Choline, 
although  by  no  means  so  poisonous  as  neurine,  has  a  similar  action 
when  administered  in  sufficiently  large  doses.  According  to  Brieger, 
it  is  about  one-tenth  to  one-twentieth  as  toxic  as  neurine.''''  Choline 
seems  to  be  rapidly  destroyed  in  the  body,  not  aj^pearing  in  the  urine  **^ 
but    forming    formic    acid    and    perhaps    glyoxylic    acid.     Donath  ''^ 

78Kutscher  and  Loliiiiaiiii.  /oil.  pliysiol.  C1u>mi..  1000    (48).  1. 

70  Halliburtxm,  "Cliciii.  of  Musclo  and  Xorvo,"  1004,  p.  110.  states  that  elioline 
produces  a  fall  in  blood  |)ressure  by  dilatinj;  tlie  porii)heral  vessels,  whereas 
neurine  constricts  tlie  periplicral  vessels;  he  uses  tliis  dillerence  in  pliysiolopical 
elTeet  as  a  means  of  distinjiuisliinLr  the  two  substances.  Injected  into  animals, 
choline  causes  a  consideralile  liut  transient  decrease  in  tlie  number  of  leucocytes 
in  tlie  blood,  followed  later  1)V  an  increase  (Werner  and  Liehtenberg,  Deut.  med. 
Woch.,   1000    (32),  22). 

80  V.  Iloesslin,  Ilofmoister's  Beitr.,  lOOG    (8),  271. 


TOXfxs  125 

fouud  that  choline  injected  directly  into  the  cortex  or  under  tlie  dura 
is  extremely  toxic,  causing  severe  tonic  and  clonic  convulsions,  and 
believes  that  choline  may  be  responsible  for  epileptic  convulsions. 
This  view  has  been  opposed,  and  properly  so,  by  Handelsmann  "^  and 
others.  The  attempt  to  ascribe  importance  to  choline  as  a  cause  of 
cither  toxic  or  therapeutic  effect  of  j"-rays  seems  also  to  be  entitled  to 
but  slight  consideration.*-  It  is  probably  a  factor  in  the  lowering  of 
blood  pressure  which  results  from  injection  of  extracts  of  various  tis- 
sues, in  which  it  is  commonly  present  in  minute  amounts,-'  for  very 
minute  amounts  of  choline  will  produce  a  decided  fall  in  blood  pres- 
sures^ 

The  Pressor  Bases. — By  decarboxylation  of  amino  acids,  amines 
are  obtained,  and  some  of  them,  notably  those  derived  from  leucine, 
tyrosine,  phenylalanine  and  liistidine,  have  a  marked  effect  on  non- 
striated  muscle.     These  are  discussed  in  Chapter  xix. 

TOXINS 

Certain  bacteria  produce  soluble  poisons  by  sj'nthetic  processes, 
which  poisons  are  secreted  into  the  surrounding  medium  and  repre- 
sent the  chief  poisonous  products  of  the  bacteria,  being  capable  of 
causing  most  or  all  of  the  symptoms  attributed  to  infection  by  the 
specific  bacteria  that  have  manufactured  them.  To  this  class  of  solu- 
ble poisons  the  term  toxin  has  now  become  limited  (for  reasons  that 
will  be  mentioned  below),  including  not  only  toxins  of  bacterial  ori- 
gin, but  also  poisons  of  similar  nature  produced  by  animals  (snake 
venoms,  eel  serum,  etc.)  and  by  plants  (ricin,  abrin,  crotin).  The 
chief  bacteria  secreting  true  toxins  are  B.  diphtherke,  B.  tetani,  B. 
■pyocyaneus,  and  B.  hotulimis  (not  including  bacteria  producing  hem- 
olytic substances  resembling  toxins).  It  will  be  seen  that  the  tenn 
toxin  has  been  greatly  narrowed  since  the  time  when  all  ptomai'ns 
and  other  poisonous  bacterial  products  were  called  toxins,  until  now 
it  ha.s  come  to  include  the  specific  poisons  of  but  four  of  the  great 
group  of  pathogenic  bacteria.*^ 

Chemical  Properties  of  Toxins. — The  chemical  nature  of  the 
toxins  is  entirely  unknown.  Hy  various  precipitation  methods  they 
may  be  carried  down,  but  included  with  them  are  masses  of  impuri- 
ties, chiefly  proteins.     They  behave  like  electro-positive  colloids,®^  but 

siZeit.  f.  physiol.  Chem.,  1003  (39),  526;  also  see  Mcd«  Xews,  100.5  (86),  107, 
for  literature  and  methods  of  analysis. 

82  See  Schenk.  Deut.  med.  Woch.',  1910    (36),  1130. 

83  Schwarz  and  Lederer,  Pfliiger's  Arch.,  1008  (124),  3.)3 ;  Kinosliita,  ihi<l..  lOlO 
(1.32),  607. 

S4  Mendel  et  al.,  Jour.  Pharm.  and  Fxp.  Ther..  1012  (3),  64S :  Hunt  and 
Taveaii,  Bulletin  73,  Hyg.  Lab.  U.  S.  P.  H.  Service. 

85  It  is  possible  that,  dysentery  bacilli,  and  perhaps  a  few  other  patliogens. 
secrete  a  small  amount  of  true  toxin.  Pick  considers  the  active  constituent  of 
tuberculin  to  be  a  true  toxin,  or  closely  related  tliereto. 

SG  Field  and  Teague,  Jour.  Exper.  Med.,  1007    (0),  86. 


126  CHEMISTRY  OF  BACTERIA  ASD  THEIR  PRODUCTS 

diffuse  faster  than  proteins.  It  is  not  certain  that  toxins  are  not 
proteins,  for  although  certain  investigators  report  that  by  purification 
processes  very  active  toxins  have  been  obtained  that  did  not  give 
the  protein  reactions,  yet  the  toxins  are  attacked  by  proteolytic  en- 
zymes, and,  like  proteins,  are  precipitated  by  nucleic  acid  (Kossel). 
Furthermore,  accumulating  experience  with  immunological  processes 
adds  increasing  doubt  as  to  the  possibility  of  antibody  formation 
being  incited  by  anything  but  proteins.  Oppenheimer  says  of  the 
toxins  that  "we  nnist  be  contented  to  assume  that  they  are  large  mo- 
lecular complexes,  probably  related  to  the  proteins,  corresponding  to 
them  in  certain  properties,  but  standing  even  nearer  to  the  equally 
mysterious  enzymes  with  whose  properties  they  show  the  most  ex- 
tended analogies  both  in  their  reactions  and  in  their  activities." 
These  similarities  between  toxins  and  enzj-mes  are  very  striking,  and 
in  discussing  the  nature  of  the  enzymes  we  have  mentioned  the  rea- 
sons for  considering  them  related  to  the  toxins ;  we  may  now  take  up 
the  other  side  of  the  question  and  consider  the  relation  of  the  toxins 
to  the  enzymes. 

Resemblance  to  Enzymes. — First  of  all  we  meet  the  same  diffi- 
culty in  isolating  toxins  that  we  do  in  isolating  enzymes.  "A  pure 
toxin  is  as  unknown  as  a  pure  enzyme"  (Oppenheimer).  At  first 
both  were  believed  to  be  proteins ;  now  both  are  considered  by  many 
not  to  be  proteins,  but  molecular  complexes  of  nearly  equally  great 
dimensions.  That  toxins,  like  enzymes,  are  colloids,  has  been  abun- 
dantly demonstrated.*"  Both  pass  through  porcelain  filters,  but  both 
lose  much  of  their  strength  in  the  process,  and  they  are  almost  en- 
tirely held  back  by  dialyzing  membranes.  They  behave  similarly  as 
regards  adsorption  by  suspensions,  and  have  similar  effects  on  the 
physical  properties  of  their  solutions  (Zunz).*-  Neither  will  with- 
stand boiling,  and  most  forms  are  destroyed  at  80°  instantly  or  in 
a  very  short  time ;  on  the  whole,  however,  toxins  are  more  susceptible 
to  heat,  as  well  as  to  most  other  injurious  agencies.  Both  stand  dry 
heat  over  100°,  and  extremely  low  temperature,  without  much  injury. 
Left  standing  in  solution  for  some  time  they  gradually  lose  their 
specific  properties,  and  in  each  case  this  seems  to  be  due  to  an  altera- 
tion in  the  portion  of  the  molecule  that  produces  the  destructive  ef- 
fects {toxophore  or  zymophore  group),  while  the  portion  of  the  mole- 
cule that  unites  with  the  substance  that  is  to  bo  attacked  (hapfophore 
group)  remains  uninjured,  the  toxin  becoming  a  ioxoid.  the  enzyme 
a  fermcntoid.  Enzymes  as  well  as  toxins  are  poisonous  when  injected 
into  animals,  and  the  animals  react  to  each  by  producing  substances 
{antihodies)  that  render  each  inert,  probably  in  the  same  way.  On 
the  other  hand,  enzymes  and  to.xins  seem  to  ])i'()(luce  their  effects  ac- 
cording to  difl'erent  laws: — A  small  amount  ol"  enzyme  can  in  course 

87  See  ZangRcr.  (Vnt.   f.   liakt.    (ivf.),   1 !)(),")    (;i(i),  2:!!). 

88  Arch,  (li   iMsiol.,    l!)(Mt    (7  ).    \'M . 


TOXINS  127 

of  time  produce  an  almost  indetiiiite  amount  of  effect,  whereas  toxins 
act  more  nearly  quantitatively.  It  seems  as  if  the  enzyme  were  bound 
to  the  body  upon  which  it  acts,  as  is  the  toxin,  but  that  after  it  has 
destroyed  this  body  it  is  set  free  in  a  still  active  form,  ready  to  ac- 
complish further  work,  whereas  the  toxin  is  either  not  set  free,  or  it 
becomes  inactive  after  it  has  once  ])een  combined. 

Agencies  Destroying  or  Modifying  Toxins. — Toxins  are  very  sus- 
ceptible to  li^lit.  direct  suidi^ht  soon  destroying  the  power  of  toxin 
solutions.  Fluorescent  sul)stances  destroy  toxins  both  in  vitro  and 
in  the  body.^''  Oxygen,  even  dilute  as  in  air,  is  harmful ;  and  all 
oxidizing  agents,  including  oxidizing  enzymes,  destroy  them  quickly."'' 
Like  enzymes,  they  withstand  such  antiseptics  as  chloroform,  toluene, 
etc.,  and  are  precipitated  by  the  heavy  metals.  Some  agencies  seem 
to  attack  only  the  toxophore  portion  of  the  molecule,  e.  g.,  iodin,  car- 
bon disulphid  (Ehrlich).  Certain  toxins  (diphtheria,  dysentery) 
can  be  converted  into  non-toxic  modifications  by  acids,  the  original 
toxicity  being  restored  by  bases  (Doerr),-'^  which  fact,  Pick  maintains, 
is  in  support  of  the  protein  nature  of  toxins.  Salts  of  monovalent 
metals  have  no  effect  on  toxins,  but  bivalent  and  trivalent  salts  are 
injurious  to  them,  tetanus  toxin  being  more  sensitive  than  diphtheria 
toxin.     X-rays  are  said  to  weaken  them."- 

Introduced  into  the  gastro-intestinal  tract,  most  bacterial  toxins 
are  not  absorbed  (botulinus  toxin  excepted),  cause  no  symptoms,  and 
do  not  reappear  in  the  feces;  they  are  therefore  destroyed  by  the 
contents  of  the  tract,  pepsin,  pancreatic  juice,  and  bile  all  being  capa- 
ble of  destroying  toxins."^  They  may,  however,  when  injected  sub- 
cutaneously,  circulate  unimpaired  in  the  blood  of  non-susceptible  an- 
imals, gradually  disappearing,  more  through  slow  processes  of  de- 
struction than  hy  elimination.  When  injected  into  susceptible  ani- 
mals, they  soon  disappear  from  the  blood,  being  fixed  in  the  organs 
that  they  attack.  Toxins  are  also  bound  hy  lipoids,  fats  and  similar 
substances,  which  accounts,  at  least  in  part,  for  the  affinity  of  tetanus 
toxin  for  nervous  tissues."*  In  common  with  other  colloids  they  are 
adsorbed  by  surfaces,  such  as  charcoal,  kaolin,  etc. ;  such  adsorption 
is  accompanied  by  little  change  in  any  of  the  physical  properties  of 
the  solution,  except  an  increase  in  surface  tension  (Zunz). 

Differences  from  Ptomains. — While  ptomains  are  formed  by  cleav- 

89  Literature  given  by  Xoguchi.  .Jour.  Exper.  ^led.,   11)06    (8),  263. 

'■'0  According  to  Pitini  (Biochem.  Zeit..  1910  (2o),  2.57)  toxins  cause  their  liarm- 
ful  effects  by  reducing  tlie  oxidizing  capacity  of  the  tissues. 

91  Wien.  klin.  Woch.,  1907    (20),  5. 

92Gerhartz,  Berl.  klin.  Woch.,   1909    (46),   1800. 

93  Baldwin  and  Levene  (.Jour.  Med.  Research,  1901  (6),  120)  found  that  diph- 
theria and  tetanus  toxin  are  both  destroyed,  apparently  tiirougli  digestion,  by 
pepsin,  trypsin  and  papain  acting  for  several  days.  Review  of  Literature  bv 
Lust,  Hofmeister's  .Beitr.,  1904  (6),  1.32.  See  Vincent,  Ann.  Inst.  Pasteur,  1908 
(22),  341. 

94Loewe,  Biochem.  Zeit.,  1911    (33),  225,  and   (34),  495. 


128  CHEMISTRY  OF  BACTERIA  AXD  THEIR  PRODUCTS 

age  processes  from  the  medium  upon  which  the  bacteria  grow,  and 
the  same  ptomains  can  be  produced  by  several  different  kinds  of  bac- 
teria, the  toxins  are  synthetic  products  of  absolutely  specific  nature. 
That  they  are  produced  by  synthesis  can  be  sho\ra  by  growing  the 
bacteria  on  Uschinsky's  or  similar  media,  which  contain  no  proteins, 
carbohydrates,  or  fats,  but  merely  simple  organic  and  inorganic  salts 
of  known  composition ;  on  these  media  the  bacteria  produce  their  spe- 
cific toxins,  which  must,  therefore,  be  synthesized.''^  Furthermore, 
diphtheria  toxin  is  essentially  the  same  no  matter  on  w^hat  sort  of 
medium  the  bacteria  are  grown,  whereas  ptomains  vary  wdth  the 
nature  of  the  substance  from  which  they  are  produced.  Toxins  are 
true  secretions  of  bacterial  cells,  just  as  tiypsin  is  of  pancreatic  cells, 
or  thyroiodin  of  thyroid  cells.  Anti-bodies  can  be  produced  against 
toxins,  but  not  against  ptomains. 

Ehrlich's  Conception  of  the  Nature  of  Toxins. — Chemical  studies 
of  toxins  being  impossible,  we  have  been  obliged  to  study  them 
through  their  physiological  effects,  just  as  we  have  obtained  informa- 
tion concerning  enzymes  through  their  specific  actions.  In  this  way 
Ehrlich  developed  well-crystallized  ideas  concerning  the  structure 
of  toxins,  as  well  as  the  manner  in  wliich  they  act,  which  may  be 
briefly  summarized  as  follows :  Each  toxin  molecule  consists  of  a  large 
number  of  organic  complexes  grouped,  as  in  other  organic  compounds, 
as  side-chains  about  a  central  chain  or  radical.  One  or  more  of  these 
complexes  has  a  chemical  affinity  for  certain  chemical  constituents  of 
the  tissues  of  susceptible  animals,  wdth  which  the  toxin  molecule 
unites;  this  binding  group  is  called  the  haptopJiore  (meaning  "bear- 
ing a  bond")-  Another  side-chain  or  group  of  side-chains  exerts  the 
injurious  effects  upon  the  tissue  to  which  the  molecule  has  been  bound 
by  the  haptophore,  and  cannot  produce  these  injurious  effects  unless 
it  has  been  so  bound.  This  injury-working  group  is  called  the  toxo- 
phore.  An  animal  is  susceptible  to  a  toxin  only  when  its  cells  con- 
tain substances  which  possess  a  chemical  affinity  for  the  haptophore 
groups  of  the  toxin,  and  also  substances  which  can  be  harmfully  influ- 
enced by  the  toxophore  groups.  Tetanus  toxin,  for  example,  ow^es  its 
effects  to  the  fact  that  nervous  tissues  contain  chemical  substances 
having  a  strong  affinity  for  the  haptophore  group  of  tetanus  toxin, 
and  also  substances  that  can  be  attacked  with  serious  results  by  the 
toxophore  group  of  the  toxin.  The  nature  of  the  changes  brought 
about  by  the  toxophore  groups  of  toxins  is  not  understood ;  there  are 
many  resemblances  to  the  action  of  enzymes,  but  the  analogy  is  by 
no  means  complete.  "We  find  perhaps  the  closest  analogy  to  the  en- 
zymes in  the  toxic  substances  that  destroy  red  corpuscles  and  bacteria 
(hemohjsi)is,  harfcrioh/si'tis) ,  wliit'li  will  be  considered  in  another 
])lace.  The  immunity  against  toxins  and  enzymes  seems  to  be  pro- 
duced by  identical  processes,  which  consist  in  an  overproduction  of 

oslladlcy,  Jour.  Infect.  Dis..  lOO",  Suppl.  TTT,  ]>.  05. 


ENDOTOXINS  129 

the  cellular  constituents  (receptors)  -which  bind  the  haptophure 
o-roups  to  the  cells,  these  excessive  receptors  being  secreted  into  the 
blood,  where  they  combine  with  the  toxin  or  enzyme  so  that  it  cannot 
enter  into  combination  with  the  cells.  This  "side  chain  theory"  of 
Ehrlich  has  been  a  useful  working  hypothesis,  although  it  is  becoming 
highly  probable  that  it  does  not  picture  the  exact  method  of  toxin  and 
antitoxin  action."^'* 

Immune  substances  cannot  1)C  produced  against  ptomdins,  or  for 
that  matter  against  the  vegetable  alkaloids,  or  against  any  chemical 
bodies  of  knoicn  constitution.  Another  difference  between  the  action 
of  toxins  and  simpler  chemical  poisons  is,  that  while  with  the  latter 
the  effects  are  produced  in  a  very  short  time  after  injection,  there  is 
a  latent  period  of  several  hours  before  symptoms  appear  after  inject- 
ing toxins.  What  occurs  during  this  latent  period  is  not  fully  known, 
but  that  there  is  a  latent  period  suggests  a  resemblance  to  enzyme 
action.  An  alkaloidal  or  other  chemical  poison  enters  the  cell,  and  its 
harm  is  done  at  once.  A  toxin  combines  with  the  cell,  and  then,  if 
it  produces  its  effects  by  an  enzymatic  alteration  of  the  cellular  struc- 
ture, some  time  must  elapse  before  the  changes  are  great  enough  to 
cause  the  appearance  of  symptoms. 

ENDOTOXINS  og 

By  far  the  greater  number  of  pathogenic  bacteria  do  not  secrete 
their  poisons  as  toxins  into  the  surrounding  medium,  although  they 
manifestly  cause  disease  by  poisoning  their  host.  Among  them  are 
such  organisms  as  the  typhoid  bacillus,  pneumococcus,  the  pus  cocci, 
cholera  vibrios,  and  many  others.  If  cultures  of  these  organisms  are 
iiltered,  the  tiltrate  will  be  found  to  be  but  slightly  toxic  (except  for 
the  hemolytic  poisons),  although  the  bodies  of  the  bacteria  after  they 
have  been  killed  by  chloroform  or  other  antiseptics  are  highly  poison- 
ous if  injected  into  an  animal.  These  bacteria,  then,  produce  poisons 
which  do  not  escape  from  the  cells  into  the  culture-medium,  but  are 
firmly  held  within  them.  By  using  various  means  these  intracellular 
toxins,  or  endotoxins,  can  be  obtained  independent  of  the  bacterial 
cells.  One  of  these  is  to  grind  up  the  cells,  which  can  be  particularly 
well  done  if  they  are  first  made  brittle  by  freezing  at  the  temperature 
of  liquid  air  (MacFadyen's  method).  By  very  great  pressure  in  the 
Buchner  press  the  cellular  contents  can  be  expressed.  They  may  also 
be  obtained  by  letting  the  bacteria  autolyze  themselves  for  a  short 
time  in  non-nutrient  fluids  (Conradi,"  et  al.)  Endotoxins  obtained 
in  this  way  are  soluble  and  highly  poisonous,  and  it  is  undoubtedly 
through  their  action  that  the  characteristic  diseases  are  produced  by 
the  bacteria  that  contain  them.     Presumably  the  endotoxins  are  liber- 

95a  See  Coca;  Jour.  Infect.  Dis.,  1915   (17),  3.51. 

96  See  general  review  by  Pfeiffer,  Jahresber.  d.  Innimnitatsforsch..  1910   (6).  13. 
97Deut.  med.  Woch.,  1903   (29),  26. 
9 


130  CHEMISTRY  OF  BACTERIA  AND  THEIR  PRODUCTS 

ated  in  the  body  either  by  autolysis,  or  by  heterolysis  by  the  enzymes 
of  the  body  cells  and  fluids,  and  there  is  some  question  as  to  whether 
they  are  preformed  specific  constituents  of  the  bacteria,  or  merely 
the  poisonous  product  of  enzymatic  disintegration  of  the  bacterial 
proteins,  similar  to  the  "  anaphylatoxins. "  "^'^ 

Endotoxins  differ  from  the  true  toxins,  however,  in  one  important 
respect:  namely,  it  is  difficult  or  impossible  to  obtain  cm  antitoxin 
for  endotoxins  by  immunization  of  animals°^  Animals  immunized 
against  endotoxins  develop  in  their  serum  substances  that  are  bacteri- 
cidal and  agglutinative  to  the  bacteria  from  Avhich  the  poisons  are 
derived,  but  the  serum  will  not  neutralize  the  endotoxins.  As  a  re- 
sult, we  are  unable  to  perform  experiments  indicating  whether  endo- 
toxins have  the  same  structure  as  the  true  toxins,  i.  e.,  a  haptophore 
and  a  toxophore  group,  but  presumably  their  nature  is  different  in 
some  essential  particular.  The  chemical  nature  of  the  endotoxins  is 
also  unknown,  for  they  are  always  obtained  mixed  with  the  other  con- 
stituents of  the  bacteria.'*'' 

Tuberculin,  once  supposed  to  be  an  albumose,  is  produced  even  when 
the  bacilli  are  grown  on  a  protein-free  medium,  and  in  the  active  solu- 
tion no  albumose  or  other  protein  is  then  found.  Hence  it  seems 
probable  that  tuberculin  is  of  the  nature  of  a  polypeptid,  which  gives 
no  biuret  reaction  but  is  destroyed  by  pepsin  and  trypsin,  according 
to  Loevenstein  and  Pick,^  but  not  by  erepsin  (Pfeiffer).-  Whether 
tuberculin  should  be  considered  an  endotoxin  liberated  by  the  disin- 
tegration of  the  bacilli  in  the  cultures  is  unknown ;  Pick  looks  upon  it  as 
a  secretion  of  the  bacilli,  and  closely  related  to  the  true  toxins. 

Since  far  more  bacterial  diseases  are  brought  about  by  endotoxins 
than  by  true  toxins,  the  failure  to  secure  antitoxins  for  these  sub- 
stances has  been  a  great  check  in  the  progress  of  serum  therapy,  and 
the  problem  of  the  endotoxins  is  one  of  the  most  important  in  the  en- 
tire field  of  immunity. 

97a  See  Dold  and  Hanau,  Zeit.  Immunitiit.,  1913  (10),  31;  Zinsser.  "Infection 
and  Resistance,"  N.  Y.,  1914,  Chap.  xvii. 

98  Positive  results  are  claimed  by  Besredka  (Ann.  Inst.  Pasteur,  1906  (20), 
304).  and  some  others;  see  Kraiis,  Wien.  klin.  Wocli..  1900  (19).  6,5,') :  Zeit. 
Immunitiit.,  1909  (3),  646.  It  is  suggested  by  VVassermann  (Kolle  and  \Yasser- 
maim's  Ilandbuch,  1912  (2),  246)  that  this  dilTiculty  in  obtaining  antiondoloxins 
depends  on  the  large  size  of  the  molecule, — the  small  diffusible  toxin  molecule  is 
so  altered  in  its  physical  condition  tlirough  union  with  the  antibody  that  its 
properties  are  much  altered,  whereas  the  large  endotoxin  molecule  must  be  di- 
gested by  com])lement  before  its  toxicity  is  destroyed. 

99 'J'lie  Afi(/7-eNsivs  of  Pail,  to  which  lie  ascrilies  the  pathogenicity  of  bacteria, 
are  too  little  esta])lished  to  permit  of  a  discussion  from  the  chemical  standpoint. 
By  many  they  are  Ixdieved  to  be  iiolhing  more  tluin  endotoxins.  (Literature 
given  by  Miilier,  Oppenhcimer'a  Ilandb.  d.  Piochem.,  1909  (II  (1)  ),  6S1  :  Dud- 
geon, Lancet,   1912    (1S2),   1673).     According  to  Ingravelle    (Ann.  d'  ig.  sperim., 

1910  (20),  483),  typhoid  aggressins  arc  found  in  the  albumins. 

1  Piwhem.  Zeit.,"  1911    (31),   142. 

2  Wien.  klin.  Woch.,  PMl    (24),  11  If);   see  also  Lockiiiaiiii.  Zeit.  pbvsiol.  C'liem.. 

1911  (73),  389. 


POISONOUS  BACTERIAL  PROTEINS  131 

POISONOUS  BACTERIAL  PROTEINS 

If  we  filter  a  bouillon  culture  of  diphtheria  bacilli  through  porce- 
lain, wash  thoroughly  with  salt  solution  the  bacteria  remaining,  and 
collect  them  thus  freed  from  their  secretion  products,  it  will  be  found 
that  extracts  of  the  bacterial  substance  or  the  bodies  of  the  killed  bac- 
teria themselves  are  quite  free  from  the  typical  toxin.  This  indicates 
that  the  toxin  is  eliminated  from  the  bacteria  as  fast  as  it  is  formed, 
and  no  considerable  quantity  is  retained  within  the  cell.  The  bac- 
terial substance,  however,  or  proteins  isolated  from  it,  is  found  to  pro- 
duce severe  local  changes  when  injected  into  the  bodies  of  animals, 
necrosis  and  a  strong  inflammatory  reaction  with  pus-formation  being 
the  chief  features.  This  local  effect  is  not  a  specific  property  of  the 
diphtheria  bacillus,  for  other  bacterial  proteins,  including  proteins 
from  non-pathogenic  bacteria,  will  produce  the  same  changes;  in- 
deed, many  proteins  that  are  derived  from  vegetable  and  animal 
sources  have  equally  marked  pyogenic  properties.  All  foreign  pro- 
teins when  introduced  into  the  circulation  of  animals  are  more  or  less 
toxic,  and  the  toxic  effects  of  the  bacterial  proteins  are,  for  the 
most  part,  neither  specific  nor  particularly  striking.  There  are  a  few 
pathogenic  organisms,  however,  which  seem  to  produce  neither  true 
toxins  nor  endotoxins,  notably  the  tubercle  bacillus  and  the  anthrax 
bacillus,  and  with  these  there  may  be  a  relation  between  their  protein 
constituents  and  their  specific  effects. 

Numerous  protein  substances  have  been  extracted  from  bacterial 
cells,  particularly  nucleoproteins,  but  also  proteins  resembling  al- 
bumins, nucleo-albumin,  and  globulins.  In  all  probability  the  chief 
proteins  of  the  bacterial  cell  are  nuelein  compounds,  wliicli  is  indi- 
cated both  by  their  nuclear  staining  and  by  the  analyses  of  Iwanoff ;  * 
and  many  of  the  nucleoproteins,  both  of  bacterial  and  non-bacterial 
origin,  cause  considerable  local  inflammatoiy  reaction  when  injected 
into  animals.  Tiberti  *  claims  that  vaccination  with  non-lethal  doses 
of  the  nucleoproteins  of  anthrax  bacilli  will  protect  animals  against 
inoculations  of  virulent  anthrax  bacilli.  Some  of  the  earlier  observa- 
tions on  the  toxicity  of  bacterial  proteins  were  erroneous  because  im- 
pure proteins,  containing  toxins,  endotoxins,  and  ptomai'ns  were  used. 
Schittenhelm  and  Weichardt  ^  have  found,  however,  that  bacterial 
proteins  are  much  more  toxic  than  any  ordinary  proteins,  as  indi- 
cated by  loss  of  nitrogen,  temperature  changes  and  alterations  in  the 
leucocytes  of  injected  animals. 

Vaughan  and  his  students  have  been  able  to  split  off  from  the 
bodies  of  various  pathogenic  bacteria  toxic  materials  which  are  stated 
to  resemble  in  some  respects  the  protamins,^^  although  they  do  not  all 

3  Hofmeister's  Beitr.,  1002    (1),  524. 

4  Cent.  f.  Bakt.,  1006    (40),  742. 
sMiinch.  med.  Woch.,  1011    (58),  841. 

5a  A  full  review  of  this  work  is  given  in  Vaucrhan's  "Protein  Split  Products," 
Philadelphia,  1013;  and  in  Jour.  Lab.  Clin.  Med.,  1916,  Vols.  1  and  2. 


132  CHEMISTRY  OF  BACTERIA  ASD  THEIR  PRODUCTS 

give  a  satisfactory  biuret  test.  These  toxic  materials  are  evidently 
quite  different  from  either  the  true  soluble  toxins  or  the  endotoxins, 
since  they  resist  heatinp:  for  ten  minutes,  at  110°  in  the  autoclave 
with  1  per  cent,  sulphuric  acid,  this  being-  a  method  used  for  securing 
the  substance.  Since  the  sarcinas  and  B.  prudigiosus  also  yield  similar 
toxic  products,  they  cannot  be  considered  as  the  specific  toxic 
substances  of  the  pathogenic  bacteria,  but  apparently  are  com- 
mon to  all  proteins  of  whatever  origin.  With  some  bacteria  the  split- 
ting process  with  sulphuric  acid  separates  completely  the  toxic  from 
the  non-toxic  insoluble  bacterial  substance,''  e.  g.,  B.  coli  communis; 
with  others  a  toxic  portion  remains  insoluble.  The  colon  bacillus  pro- 
tein gives  all  the  protein  reactions,  is  synthesized  on  Uschinsky's 
medium,  and  does  not  yield  a  reducing  carbohydrate.  From  B. 
fifphosiis  about  10  per  cent,  by  weight  of  protein  can  be  split  off  by 
dilute  acid,  of  which  at  least  a  part  seems  to  be  a  phospliorized  glyco- 
protein.'^ Poisonous  substances  have  also  been  obtained  from  B.  diph- 
therice,  B.  anthracis,  B.  tuherculosis  ^  and  B.  pyocyaneus.  They  pro- 
duce death  without  the  usual  latent  period  obsen'ed  with  toxins,  and 
are  very  toxic,  a  few  (10-20)  milligrams  of  colon  bacillus  poison  kill- 
ing guinea-pigs  in  less  than  ten  minutes.''  A  certain  degree  of  immu- 
nity can  be  obtained  against  them.^°  Their  relation  to  endotoxins  has 
yet  to  be  determined. 

BACTERIAL  PIGMENTS  i^ 

The  formation  of  pigment  by  bacteria  seems  to  be,  for  the  most 
part,  an  adventitious,  unessential  property.  There  are  a  few  bacteria 
wliicli  possess  pigments  of  the  nature  of  chlorophyll,  or  allied  to  it, 
and  this  pigment  is  undoubtedly  of  great  importance  in  the  life 
processes  of  these  particular  forms.  Other  varieties  of  pigment- 
forming  bacteria,  of  which  but  very  few  are  pathogenic  {Bacillus 
pyocyaneus,  B.  proteus  fluorescens,  S.  pyogenes  aureus  and  citreus, 
M.  cereus  flavus),  seem  to  produce  pigment  as  a  waste  product  which 
is  excreted  from  the  cell  as  fast  as  formed.  Generally  the  pigments 
are  produced  in  a  colorless  form  (leuco-hase)  which  is  oxidized  by  the 
air  into  the  pigment,  e.  g.,  in  pyocj/aneus  infections  the  soiled  dress- 
ings are  most  colored  about  the  portions  most  exposed  to  air.  Since 
pigment-forming  bacteria  produce  pigments  only  under  certain  condi- 
tions, and  can  grow  abundantly  without  producing  any  pigment,  it 
is  evident  that  the  pigment  formation  is  no  very  essential  part  of 
llicif   iiK'tjiholisiii.     It   is   ])()ssi])]('  to  modify   pigment   ])r()ducti()ii    al- 

G  Wlioelcr,  Jour.  Amor.  ^Tcd.  Assoc,  1905    (44),  1271. 
■!  Ihid.,  in04    (42).  1000. 

«S(>o  White  and  Avery,  Jour.  IVfed.  Pxes.,  1912   (20),  317. 

0  Jour.  Amcr.  ]\Ied.  Assoe..  lOOfi   (44),  1.340;  American  Medic  iiie.  inO.l   (10).  14"!. 
lOVaufrhan    (Jr.),  Jour,  of  INIed.  riesearch,  1905    (14),  ()7. 

13  For  complete  bibliography  and  r^surn^  see  Sullivan.  Jour.  ^led.  Research, 
1905    (14),   109. 


BACTERIAL  PIGMENTS  133 

most  at  will,  and  even  to  develop  races  of  bacteria  that  do  not  produce 
pigment  at  all,  from  races  that  ordinarily  are  pigment-producers. 

Of  nuniorous  classifications  of  pigment-forming  bacteria,  all  faulty 
because  of  our  slight  knowledge  of  the  chemistry  of  the  process,  that 
of  Migula  seems  the  best;  it  is  based  on  the  solubility  of  the  pigments 
formed,  as  follows: 

(1)  Pig-ments  Soluble  in  Water. — This  includes  the  pigments  of 
all  fluorescent  bacteria,  as  well  as  those  giving  a  red  or  brown  color 
to  gelatin  media.  Most  important  among  these  is  Bacillus  pyo- 
cyaneus,  whose  pigments  have  been  considerably  studied.  There  seem 
to  be  two  pigments,  one,  pyocyanin,  characteristic  for  this  organism; 
and  a  fluorescent  pigment  which  numerous  other  organisms  also  pro- 
duce. Pyocyanin  has  been  analyzed  by  Ledderhose,  who  found  it  to  be 
a  ptoma'in-like  body,  a  derivative  of  the  aromatic  series,  probably  re- 
lated to  the  anthracenes.  It  can  be  reduced  to  a  colorless  leuco-base,  in 
which  form  it  is  probably  produced  by  the  bacteria,  and  then  is 
oxidized  in  the  air  into  the  pigment.  Its  composition  is  Cj^Hj^NoO 
(the  sulphur-containing  pyocyanin  which  has  been  described  is  proba- 
bly impure).  The  fluorescent  pigment  is  insoluble  in  alcohol  and 
chloroform,  and  can  thus  be  separated  from  pyocyanin,  which  is  solu- 
ble in  chloroform.  Although  related  to  the  ptomains,  pyocyanin  seems 
to  be  altogether  non-poisonous  to  animals. 

Jordan  ^*  and  Sullivan  ^^  have  studied  the  conditions  under  which 
pigments  are  formed,  and  found  that  pyocyanin  can  be  produced  in 
protein-free  media,  and  without  the  presence  of  either  phosphates  or 
sulphates;  but  both  sulphur  and  phosphorus  must  be  present  to  pro- 
duce the  fluorescent  pigment.  As  pigments  can  be  produced  on  media 
containing  only  ammonium  salts  of  succinic,  lactic,  or  aspartic  acid, 
or  asparagin,  they  are  evidently  formed  synthetically,  and  not  by 
cleavage  of  the  media. 

(2)  Pigments  Soluble  in  Alcohol  and  Insoluble  in  Water. — The  most 
important  bacteria  of  this  group  are  the  Staphylococcus  pyogenes 
aureus  and  citreus.  Their  pigment  is  of  a  fatty  nature,  a  lipochrome, 
which  lies  among  the  bacteria  in  the  form  of  dendritic  crystals.  Be- 
ing a  fat,  it  can  be  saponified,  and  when  decomposed  it  gives  the 
acrolein  reactions  and  odor,  from  the  breaking  down  of  the  glycerol 
of  the  fat  molecule.  Acted  upon  by  strong  sulphuric  acid,  the  yel- 
low pigment  changes  into  blue  granules  and  crystals  {lipocyanin  re- 
action). The  lipochromes  are  soluble  in  the  usual  fat  solvents,  and 
fonn  fat  spots  on  paper. 

(3)  Pigments  Insoluble  in  Water  and  in  Alcohol. — The  pigment  of 
Micrococcus  cereus  flavus  belongs  to  this  class;  its  nature  is  quite  un- 
known. 

i*Jour.  Exper.  Med.,   1899    (4),  627. 


CHAPTER    V 
CHEMISTRY  OF  THE  ANIMAL  PARASITES  ' 

This  subject  has  received  muck  less  consideration  than  its  import- 
ance deserves,  and  we  are  quite  in  the  dark  as  to  how  much  of  the 
effects  produced  by  animal  parasites  are  not  merely  mechanical,  but 
are  due  to  soluble  poisons  that  they  may  secrete  or  excrete.  Some  of 
the  parasites  probabl.y  cause  harm  mechanically  and  in  no  other  way, 
but  with  most  of  them  there  is  more  or  less  evidence  of  the  forma- 
tion of  poisonous  substances.  The  composition  of  the  bodies  of  the 
animal  parasites  is  an  almost  unexplored  field,  but  we  have  no  reason 
to  believe  that  the  composition  of  the  cells  of  invertebrates  differs 
essentially  from  that  of  the  cells  of  higher  organisms.  Perhaps  the 
most  characteristic  constituent  observed  in  many  forms  is  chit  in,  which 
forms  a  large  part  of  the  outer  covering  of  the  encysted  forms,  and 
probably  of  many  of  the  worms.  Glycogen  is  usually  abundant  in 
the  invertebrates,  and  the  animal  parasites  form  no  exception,-  this 
carbohydrate  having  been  found  in  their  bodies  by  many  observers. 

Eosinophilia. — One  of  the  most  characteristic  features  of  the 
animal  parasites  is  that  they  exert  a  positive  chemotaxis,  particularly 
for  eosinophile  leucocytes.^ 

An  increase  in  the  number  of  these  cells  in  the  blood,  as  well  as 
a  local  accumulation  in  the  tissues  nearest  the  parasite,  has  been 
observed  in  infection  with  practically  all  the  animal  parasites.*  Of 
these,  infection  with  Trichinella  spiralis  causes  the  most  pronounced 
eosinophilia,  presumably  because  of  the  great  number  of  parasites 
present  in  the  tissues  at  once.  That  the  eosinophilia  is  due  to  the  ac- 
tion of  the  soluble  products  or  constituents  of  the  parasites  has  been 
shown  by  experimental  injection  into  animals  of  extracts  from  the 
bodies  of  the  parasites.  Calamida  lias  found  that  extracts  of  dog 
tapeworms  also,  when  placed  in  the  tissues  in  a  capillary  tube,  cause 
an  accumulation  of  eosinophile  cells  in  the  tube.^'  Experimental  in- 
fection with  excessive  numbers  of  trichinella  causes  a  rapid  diminu- 

1  General  references  to  this  subject  will  be  found  in  v.  Fiirtli's  "Verirli'ichende 
chemische  Physiolofjie  dor  niederen  Tiere."  .Tena,  190.3;  Faust's  "Tierische  Gifte."' 
Braunscliweip,   100(»;   Kocb.  Er<;ebnisse  Pathol..   1010    (xiv    (1)   ),  41. 

2  See  Pniifrer,  Pniiper's  Arch..  190:?    (OH),  l.l.'^ 

3  Mtorature  bv  Oi)ie.  Amer.  .Tour.  Med.  Sci.,  1904  (127).  477:  Rtiiubli.  Deut. 
Arch.  klin.  Med."  1906  (S.5),  280;  Hubner,  ihid.,  1911  (104),  2HG;  Schwarz.  Frgeb. 
allfr.  Pathol..   1914    (17,).   1.38. 

4  Litcratino  by  Pruns.  Liefmann  and  Miickel,  Miinch.  nied.  Wocli..  190,")  (ri2), 
253:  Vallillo,  Arch.  wiss.  u.  prakt.  Tierhk.,  1908    (34),  50.'). 

0  Nefrative  results  were  obtained  with  extracts  of  flclrrostoiiia  cipniiiiu)  by 
Grosso   (Folia  llcniatol.,  1912    (14),  18). 

134 


PROTOZOA  135 

tiou  iu  tlie  number  of  eosinophile  leucocytes,  Avhicli  also  show  evi- 
dences of  disintegration  in  the  bone-marrow  and  lymph-glands.  Such 
large  injections  are  fatal,  which  suggests  that  the  eosinophilia  has  a 
protective  influence.  In  favor  of  this  view  is  the  observation  of 
Milian,*^  who  found  that  sarcosporidia  in  beef  are  destroyed  by  a 
violent  leucocytic  reaction,  the  prevailing  cell  being  the  eosinophile. 
As  the  eosinophile  increase  does  not  occur  until  several  days  after 
the  infected  flesh  is  eaten,  the  chemotactic  substance  is  not  liberated 
from  the  encapsulated  trichinella?  when  tlieir  capsules  are  digested 
ofit"  in  the  gastric  juice,  but  comes  either  from  the  free  larvae,  or  from 
the  degenerated  muscles  in  which  they  burrow.  Coincident  bacterial 
infection  may  reduce  the  number  of  eosinophiles.  Herrick "  finds 
that  extracts  of  Ascaris  himhricoides  cause  a  notable  eosinophilia,  but 
only  when  the  animal  has  been  sensitized  previously  with  the  same 
extract,  the  active  agent  of  which  is  a  protein ;  this  suggests  a  rela- 
tionship between  parasitic  and  anaphylactic  eosinophilia.'^^  That  the 
eosin()j)hiles  play  a  part  in  the  immunity  reactions  obsei"\'ed  in  the 
hosts  of  animal  parasites  is  indicated  by  the  fact  that  hydatid  fluid 
loses  its  antigenic  properties  when  in  contact  with  eosinophiles.'*^ 

PROTOZOA 

These  unicellular  forms  possess  all  the  chemical  characters  of  the 
(^ells  of  higher  forms,  even  to  the  more  specialized  constituents.  Thus 
it  has  been  demonstrated  that  protozoa  contain  proteolj^tic  enzymes,^ 
and  that  they  secrete  an  acid  into  their  digestive  vacuoles.^  On  the 
other  hand,  Amcfha  coll  does  not  seem  to  digest  the  red  corpuscles  and 
the  bacteria  that  it  takes  up.^°  Whether  the  Anmha  coli  produces 
any  toxic  materials,  specific  or  non-specific,  has  not  yet  been  deter- 
mined, but  the  necrosis  that  it  produces  in  liver  abscesses,  when  bac- 
terial cooperation  can  often  be  excluded  by  culture,  strongly  indi- 
cates the  production  of  necrogenic  substances.  Apparently  these  sub- 
stances are  not  chemotactic,  in  view  of  the  absence  of  leucocytic  ac- 
cumulation which  is  characteristic  of  the  lesions  of  amebic  dj^sentery. 
There  is  also  no  evidence,  clinical  or  experimental,  that  amebic  in- 
fection causes  the  formation  of  anti-substances  of  any  kind  in  the 
body  of  the  host.  The  spontaneous  recovery  from  amebic  and  other 
protozoan  infections,  however,  may  be  considered  as  indicating  the 
development  of  an  immunity  against  these  organisms.^^     Numerous 

6  Bull,  et  Mem.  Soc.  Anat..  1901   (Ser.  G,  T.  3i,  32.3. 

7  Arch.  Int.  Med.,  1913   (11),  165. 

Ta  Supported  liv  Paulian,   Tresse  IMed..    191.1    (23),  403. 

■b  Weinberg'  and  Sepuin.  Ann.  Inst.  Pasteur.   1916    (30),  323. 

sMouton.  Conipt.  Rend.  Soc.  Biol.,  1901    (.13),  801. 

9  Le  Dantec,  Ann.  Inst.  Pasteur,  1890  (4),  776;  Greenwood  and  Saunders,  .Tour, 
of  Phvsiol.,  1894   (16),  441. 

Jo^fusgrave  and  Clegg,  Bureau  of  Gov't.  Laboratories,  ^lanila,  1904,  Xo.  18, 
p.  38. 

11  Concerninfr  immunitv  to  protozoan  infections  see  Schilling-,  Kolle  and  Wasser- 
mann's  Handbuch,   1913 '(7),  566. 


136  CnEMISTBY    OF    THE    AXIMAL    PARASITES 

observers  have  suggestecl  the  possil)ility  of  obtaining  artificial  im- 
munity against  protozoa,  and  Rossle  ^-  has  obtained  immune  sera 
against  infusoria. 

The  serum  of  rabbits  immunized  against  amoebae  was  found  by 
Sellards  ^^  to  be  cytolytic  for  the  same  amoebae,  but  no  antibodies 
could  be  found  in  the  blood  of  patients  with  amebic  dysentery. 
Novy  ^*  has  obtained  immunity  against  trypanosomes,  but  the  serum 
of  immune  animals  will  not  confer  passive  immunity.  Braun  and 
Teichmann,^^  however,  claim  positive  results  with  immune  serum 
from  rabbits;  they  found  no  poisonous  agent  in  trypanosome  sub- 
stance.^^" The  fact  that  trypanosomes  themselves  readily  become  im- 
mune to  various  trypanocidal  chemicals  has  been  demonstrated  and 
extensively  studied  in  Ehrlich's  laboratory.  Gonder  ^"^  has  made  the 
niteresting  observation  that  trypanosomes  which  can  be  stained  by 
certain  vital  stains,  become  unstainable  while  alive  if  immune  to 
arsenic  compounds,  suggesting  that  this  immunity  is  associated  with 
considerable  structural  or  chemical  changes. 

Plasmodium  malarise  undoubtedly  produces  toxic  substances,  which 
seem  to  be  of  such  a  nature  that  they  do  not  diffuse  from  the  red 
corpuscle,  but  are  only  liberated  when  the  corpuscle  breaks  up  on 
the  maturation  of  the  parasite.  In  this  way  the  characteristic  par- 
oxysmal manifestations  of  the  disease  are  produced.  The  nature  of 
the  poison  or  poisons  is  unknown,  but  we  have  evidence  that  it  is 
hemolytic,  since  malarial  serum  may  hemolyze  normal  corpuscles,^" 
and  extracts  of  the  parasites  are  strongly  hemolytic  (Brem^®)  ;  prob- 
ably the  malarial  hemoglobinuria  is  caused  by  this  hemolysis.  Pre- 
sumably malarial  poisons  are  not  extremely  toxic  for  parenchymatous 
cells,  since  the  parenchymatous  lesions  in  malaria  seem  to  be  relatively 
slight  as  compared  with  the  intensity  and  duration  of  the  intoxication. 
Some  authors  state  that  the  toxicity  of  the  urine  is  increased  after 
the  paroxysm,^**  which,  however,  does  not  necessarily  indicate  that 
a  poison  formed  by  the  parasites  is  excreted  in  the  urine.  Immunity 
seems  to  be  seldom  developed  against  the  malarial  poison  or  against 
the  parasite  itself,  although  some  persons  seem  to  be  naturally  im- 

12  Arch.  f.  Hvfj.,  1905    (54),  1;   full  review  of  this  topic. 

13  Philippine" Jour.  Sci.,  1911    (6).  281. 

14  Jour.  Infec.  Dis.,  1912    (11).  411. 

isZcit.  Immunitiit.,  Ref.,  1912    (6),  4G5.  • 

isallintze  (Zeit.  f.  Hyp:.,  1915  (80),  .377)  obtained  little  immunity  with  T. 
hrucei.  hut  Schillinfj  and  Rondoni  (Zeit.  Immunitiit.,  1913  (18),  651)  obtained  a 
yxiison  from  Napana  trypanosomes  which  produced  active  immunity  in  mice. 
When  trypanosomes  are  killed  by  weak  electric  currents  they  may  liberate  an 
active  poison    (Uhlenhuth  and  Sevderhelm,  Zeit.  Tmiminitiit.,   1914    (21),  30()). 

10  Zeit.  Immunitiit.,   1913    (15),"  257. 

17  See  Repnault,  Revue  de  IVIed..  1903    (23),  729. 

18  Arch.  Int.  Med.,   1912    (9),   129. 

19  Quoted  from  Blanchard,  Arch.  d.  Parasitol.,  1905  (10),  83;  this  article  pives 
a  resume  of  the  subject  of  the  toxic  substances  produced  by  the  animal  parasites. 


CKHTOliEK  l^*^ 


mune    while   some    acquire    innnunity    through    previous   nifection.- 
The  blood  of  persons  with  malaria  seems  to  contain  no  antibodies  tor 
the  parasite  (Ferrannini)r^  although  it  seems  to  have  some  antihemo- 
lytic  power   (Brem).     (Concerning  the  pigment  present  m  the  ma- 
larial parasites  see  "Pigmentation,"  Chap,  xvi.)  ,      ,     ,,    ^ 
Sarcosporidia  of  sheep  yield  aqueous  and  glycerol  extracts  that 
are  hiohlv  toxic  for  rabbits   (Pfeiffer),  the  poisonous  constituent  of 
which %as  called  sarcocusiin  by  Laveran  and  ^lesnil.-     Tins  is  so 
highlv  toxic  that  0.0001  gm.  is  fatal  to  rabbits  (per  kilo),  other  ani- 
mals "being  less  susceptible.     It  loses  its  toxicity  on  heating  at  80 
for  twentv  minutes,  and  is  impaired  at  55-57°   for  two  hours      It 
produces  pruritis  and  other  anaphylactic  symptoms,  and  although  the 
serum  of  sheep  with  this  parasite  does  not  confer  passive  anaphylaxis 
to  sarcosporidia,  vet  it  does  give  positive  complement  fixation.--     Ihat 
it  is  a  true  toxin  is  shou^n  by  Teichmann  and  Braun,-^  who  produced 
an  effective  antitoxin  by  immunizing  rabbits;  only  rabbits  seem  to 
be  susceptible  to  the  toxin.     The  sarcosporidia  contain  also  a  distinct 
theinnostable  agglutinin.     The  lethal  dose  of  dried  substance  of  sar- 
cosporidia is.  for  rabbits,  but  0.0002  gm.,  and  the  poison  seems  to 
unite  with  the  lipoids  of  the  nervous  system   (Teichmann).-      it  is 
probable  that  the  pathogenic  protozoa,   at  least  in  some  instances 
have  a  semipermeable  membrane  about  them,  for  Goebel  -^  found  that 
trypanosomes  are  very  susceptible  to  changes  in  osmotic  conditions. 

CESTODES 
T^nia  echinococcus  has  been  by  far  the  most  studied,  its  abundant 
fluid  content  furnishing  suitable  material  for  investigation.  That 
this  fluid  is  toxic  has  been  repeatedly  observed  when,  through  rup- 
ture or  puncture,  the  fluid  has  escaped  into  the  body  cavities;  such 
accidents  are  aften  followed  by  violent  intoxication,  sometimes  by 
death  ="  As  long  as  the  cvst  is  unopened  no  toxic  manifestations  are 
observed  The  most  constant  symptoms  are  local  irritation  and  in- 
flammation, accompanied  by  urticaria,  which  may  also  be  produced 
experimentally  in  man  if  the  cyst  contents  are  injected  subcutane- 

""""The  svmptoms  are  so  strikingly  similar  to  those  of  anaphylactic 
intoxication,  that  it  is  now  generally  believed  that  they  are  the  result 
of  such  a  reaction  in  a  pei^son  sensitized  by  absorption  of  antigenic 
substances  from  the  cyst."     Carriers  of  echinococcus  cysts  have  been 

20  See  Celli.  Cent.  f.  Bakt..  1900   (27).  107. 

2iRiformaMed..  1911    (27),  177. 

22Compt.  Rend.  Soc.  Biol..  1800    (51).  .311. 

22a  McGowan.  .Tour.  Path,  and  Bact.,  1913    (18),  \lo. 

24  7b7rf  '  l01o7''0)*'."96:  see  also  Knebel.  Cent.  f.  Bakt.,  1912    (66),  52.3. 
25Ann.*Soe.  Med.  d.  le  Gand.  1900   (86).  11. 
26Rpp  4ehard    \rch    "en.  de  Med..  1887    (22),  410  and  5(2. 

27!::  BoS^in  and  Lar'-oche.  Presse  Med.,  1910  (18),  329;  Ghedini  and  Zamorani, 
Cent.  f.  Bakt.,  1910  (55),  49. 


138  CHEMISTRY    OF    THE    ANIMAL    PARASITES 

found  to  have  in  their  blood  antibodies  giving  precipitin  ^^  and  com- 
plement fixation  -"  reactions  with  extracts  of  echiuococcus,  and  some- 
times with  other  taenia.^"  The  antigen  of  the  echinococcus  is  be- 
lieved by  some  to  be  a  lipoid  ;^^  in  the  case  of  Taenia  sagitiata,  at 
least,  it  seems  to  be  associated  with  the  lecithin  (Meyer  ^").  Graetz,^^ 
however,  states  that  the  protein  of  the  hydatid  cyst  is  derived  from 
the  host,  and  that  it  is  therefore  incapable  of  causing  anaphylaxis  in 
that  host,  but  it  may  undergo  alterations  in  the  cyst  so  that  it  is 
toxic  after  the  order  of  anaphylatoxins  {q.  v.).  The  complement  fixa- 
tion reaction  with  echiuococcus  fluid  has  been  found  quite  reliable  in 
the  clinic,  93  per  cent,  of  positive  reactions  having  been  obtained  in 
500  cases  collected  by  Zapelloni,^-''  while  controls  were  always  negative. 

The  fluid  of  the  echinococcus  cysts  has  generally  a  specific  gravity 
of  1005-1015,  and  contains  1.4-2  per  cent,  of  solids.  Most  abundant 
are  sodium  chloride,  about  0.8  per  cent.,  and  sugar,  0.25  per  cent., 
the  latter  presumably  coming  from  the  glycogen  contained  in  the 
wall.  Cholesterol  is  often  abundant,  while  inosite,  creatin,  and  suc- 
cinic acid  are  often  found.  Clerc  has  found  traces  of  lipase,  but 
other  enzymes  seem  to  be  absent  or  in  very  small  amounts.  Proteins 
are  present  only  in  traces,  unless  inflammation  has  occurred.  Schil- 
ling ^^  found  the  molecular  concentration  of  the  cyst  fluid  to  be  quite 
the  same  as  that  of  the  patient's  blood.  The  fluid  is  said  not  to  be 
toxic  to  laboratory  animals.^* 

The  cyst  wall  consists  of  a  hyaline  substance  which  seems  to  stand 
between  the  chitin  and  the  proteins,  and  probably  consists  of  a  mix- 
ture of  both.  Because  of  the  chitin  it  yields  about  50  per  cent,  of 
a  reducing,  sugar-like  body  when  boiled  with  acid.  Glycogen  is  also 
usually  present,  but  it  is  limited  to  the  germinating  membrane.^^ 

Other  cestodes,  when  in  the  cystic  form,  contain  fluids  which  are 
more  or  less  toxic.  Thus  Moursou  and  Schlagdenhauffen  ^^  found  a 
"leucoma'in''  in  the  Cysticercus  tenuicollis,  the  larva  of  Taenia  mar- 
ginata,  which  causes  urticaria  and  other  toxic  symptoms  when  in- 
jected into  animals.  The  fluids  of  Cysticercus  pisiformis  (the  com- 
mon cestode  of  rabbits)  have  been  found  toxic  for  frogs,  and  Vaulle- 
geard  ^^  has  determined  the  presence  of  an  "alkaloid"  and  a  ''fer- 
ment toxin"  in  this  fluid.  The  fluids  of  the  cysts  of  Caenurus  cere- 
hralis,  Coenurus  serialis,  and  Echinococcus  polymorphiis  have  all  been 

28  Welch,  et  al.,  Lancet,  1009,  Apr.  17. 

29Kreuter,  Miinoh.  mcd.  Woch.,  1900  (50),  1828;  Weinberg,  Ann.  Inst.  Pasteur 
1909    (2.3),  472. 

3o\Ii.v(.r,  Berl.  klin.  Woch.,  1910   (47).  1310;  Zeit.  Inmiunitjit.,  1910  (7).  732. 

31  Israel,  Zeit.  ITyg.,  1910  (00),  487;  Meyer,  Zeit.  Immunitiit..  1911    (9),  530. 

32  Zeit.   Iiniiumiliit.,   19)2    (15),  GO:   general  review. 
32aPoliclinico,  Surg.,  1915    (22),  Nos.  0-11. 

33  Cent.  inn.  Med.,   1904    (25),  833. 

340raet/„  Cent.  f.  Hakt.,   1910   (55),  234;   ZiMt.   IniimiMitiit ..   1912    (15),  00. 
85  Brault  and  Loeper,  Jour.  Phys.  et  Patli.  g^n.,  1904    (0),  295. 
soCompt.  Rend.  Soc.  Riol.,  1882    (95),  791. 
37  Bull.  Soc.  linneenne  de  Normandie,  1901    (4),  84. 


CESTODES  139 

found  toxic,  and  it  is  probable  that  this  is  a  general  rule  with  the 
cestodes,^^  but  human  forms  other  than  the  echinoeoccus  seem  not  to 
have  been  investigated ;  ^^  according  to  Jammes  and  Mandoul,  extracts 
of  taniia  are  bactericidal.^" 

Dibothriocephalus  latus  frequently  causes  anemia,  which  has  been 
attributed  to  a  poison  liberated  by  the  parasite  when  it  undergoes 
disintegration,  and  possibly  as  a  secretion  of  the  living  worm.*^  All 
the  intestinal  cestodes  are  equipped  with  a  well-developed  excretoiy 
apparatus,  and  it  is  easy  to  imagine  that  their  excretory  products 
may  be  toxic  to  the  animal  into  whose  intestine  they  are  excreted. 
Tallqvist  ^-  has  made  extensive  studies  of  bothriocephalus,  which  show 
that  the  active  hemolytic  agent  is  contained  in  the  lipoids  of  the 
parasites,  presumably  as  a  cholesterol  ester  of  oleic  acid.^^  The 
proglottides  contain  a  proteolytic  enzyme,  which  apparently  digests 
the  substance  of  dead  segments,  liberating  the  hemolytic  lipoid,  which 
constitutes  about  ten  per  cent,  of  the  solids  of  the  parasite.  There 
is  also  a  hemagglutinin,  which,  unlike  the  hemolytic  substance,  is 
thermolabile,  and  causes  the  appearance  of  an  antibody  in  immunized 
animals.  In  common  with  other  parasites,  antitrj^jtic  and  antipeptic 
effects  are  exhibited  by  extracts. 

Rosenqvist  **  has  studied  the  metabolism  of  twenty-one  cases  of 
bothriocephalus  anemia,  and  found  evidence  in  nearly  all  of  a  toxo- 
genic  destruction  of  protein,  which  ceases  promptly  when  the  worms 
are  removed.  He  has  found  that  these  worms  produce  a  poison 
which  is  globulicidal,  and  probably  also  generally  cytotoxic,  since  in 
the  anemias  that  they  produce,  the  elimination  of  purine  bodies  of 
tissue  origin  (endogenous  purine)  is  increased.  The  nitrogenous 
metabolism  is  quite  the  same  in  pernicious  anemia  and  in  bothrio- 
cephalus anemia.  Isaac  and  v.  d.  Velden  ^^  state  that  the  blood  of 
patients  infected  with  this  parasite  gives  a  precipitin  reaction  with 
autolytic  fluid  obtained  from  bothriocephalus,  and  that  rabbits  im- 
munized with  such  autolytic  fluids  developed  a  precipitin. 

Other  Taenia. — There  is  much  less  evidence  that  other  forms  of 
taenia  produce  toxic  substances  which  injure  their  host,  although  the 
clinical  manifestations  observed  in  persons  harboring  ta?nia  are  often 
of  such  a  nature  as  to  indicate  strongly  an  intoxication.  Jammes 
and  IMandoul  *®  found  no  toxic  manifestations  produced  by  extracts  of 
Taenia  saginata,  which  negative  finding  is  supported  by  Cao,"*"  Tall- 

38  Blanchard.,  loc  cit. 

•^n  vSemaine  med.,  190.5   (25),  55. 

•10  See  also  Joyeux,  Arch.  d.  Parasitol.,   1007    (11),  400. 

41  Literature  by  Blanchard,  Joe  rif. 

42Zeit.  klin.  iVied.,  1007    (61),  427. 

43  Faust  and  Tallqvist,  Arch.  exp.  Path.  u.  Pharm.,  1007    (57),  .307 

44Zeit.  klin.  Med.,  100,3    (40),  103. 

■*->  Dout.  med.  Woeh.,  1904    (30),  082. 

4fi  Compt.  Rend.  Acad.  Sci.,  1904    (138),   1734. 

47Riforma  med.,  1901    (3),  795. 


140  CIIEMTFiTRY    OF    THE    AMMAL    PARASITES 

qvist  and  Boycott/**  using  various  sorts  of  tienia.  These  results  con- 
tradict the  earlier  positive  finding's  of  Messineo  and  Calamida/''  who 
found  extracts  of  taenia  from  dog's  to  be  hemolytic,  chemotactic 
(especially  for  eosinophilos),  and  to  cause  local  fatty  degeneration 
in  the  liver.  Extracts  of  2\  perfoliata  and  plicata  (of  the  horse) 
were  found  highly  toxic  for  guinea-pigs  by  Pomella,'^"  the  hema- 
topoietic organs  being  greatly  stimulated.  Bedson  ^"^  found  that  ex- 
tracts of  all  sorts  of  helminths  produced  similar  effects  on  guinea-pigs, 
the  chief  lesions  being  in  the  adrenals  and  thyroid.  Possibly  these  dif- 
ferences in  results  are  due  to  the  fact  that  different  parasites  were 
studied  by  different  investigators;  furthermore,  tests  of  toxicity  of 
human  parasites  upon  rabbits  and  guinea-pigs  can  hardly  be  consid- 
ered conclusive.  Le  Dantec  did  not  find  a  precipitin  for  Taenia 
saginata  extracts  in  the  blood  of  persons  harboring  this  parasite,  and 
negative  results  with  several  other  taenia  were  obtained  by  Langer,^^ 
but  complement  fixation  reactions  may  be  given. ^^ 

Picou  and  Ramond  °^  state  that  tienia  extracts  undergo  putrefaction 
very  slowly,  and  attribute  this  to  a  bactericidal  property,  which  was 
observed  with  several  forms  of  t^nia  by  Allesandrini.  AVeinland  '"* 
has  found  that  many  intestinal  parasites  exhibit  antitryptic  proper- 
ties,^^' but  in  a  study  of  the  histological  changes  of  autolysis  I  observed 
a  taenia  in  the  intestine  of  a  dog  undergo  more  rapid  karyolytic  changes 
than  did  the  intestinal  epithelium.  Dastre  and  Stessano  ^"^  state  that 
extracts  of  Taenia  serrata  act  upon  enterokinase  rather  than  on  tryp- 
sinogen. 

NEMATODES 

Ascaris. — The  toxicity  of  members  of  this  group  has  been  a  matter 
of  dispute,  although,  as  with  the  Taenia,  there  have  been  observed  in 
patients  symptoms  that  were  more  easily  explained  as  due  to  chemical 
substances  than  as  due  to  mechanical  irritation.  INIiram,  while  study- 
ing Ascaris  megalocephala,  suffered  from  attacks  of  sneezing,  lachry- 
mation,  itching,  and  swelling  of  the  fingers,  v.  Linstow  suffered  from 
a  severe  attack  of  conjunctivitis  with  chemosis  after  touching  his  eye 
with  a  finger  that  had  been  in  contact  with  one  of  these  worms. 
Others  have  had  similar  experiences,  and  it  has  been  found  that  the 
fluid  from  these  worms  is  toxic  to  rabbits.     In  man  it  seems  to  affect 

48  Jour.  Pathol,  ajid  Bacteriol.,  1905    (10),  383. 

49  Cent.  f.  Bakt.,  1001    (;iO),  346  and  374. 
•"'OCompt.  Rend.  Soc.  Biol.,  1912    (73),  445. 
50a  Ann.  Inst.  Pasteur,  1913    (27),  682. 

•"•i  Munch,  med.  Wocli.,  1905    (52),  1665. 
•'••2Mever,  Zeit.  Imniuuitiit.,   1910    (7),  732. 
saCompt.  Rend.  Soe.  Biol.,  1899    (51),  176. 
MZeit.  f.  Biol..  1902    (44),  1  and  45. 

5fj  ('orrol)orated  for  Taenia  sa<finata  by  l-'oltorolf  (Univ.  of  Poinisvlvania  ]\Iod. 
Bull..   1907    (20),  94). 

5«  Compt.  Rend.  Soc.  Biol.,  1903   (55).,  130. 


NEMATODES  141 

especially  those  who  have  been  sensitized  by  previous  poisoninj^,  some 
persons  being  entirely  insusceptible. 

An  extensive  investigation  of  ascaris  from  both  the  chemical  and 
toxicological  standpoint  has  been  made  by  Flury,"  which  indicates 
the  source  and  nature  of  these  toxic  substances.  Because  of  the 
practically  anaerobic  conditions  under  which  the  worms  live,  Flury 
believes,  tlie  products  of  their  metabolism  are  characterized  by  being 
incompletely  oxidized,  and  resemble  the  products  of  anaerobic  bac- 
teria. Most  important  of  these  are  volatile  aldehydes  and  fatty  acids, 
especially  valerianic  and  butyric  acids,  in  less  quantities  formic, 
acrylic  and  propionic  acids.  The  toxicologic  action  of  these  volatile 
substances  is  of  such  a  character  as  fully  to  explain  the  severe  irrita- 
tion of  skin  and  mucous  membranes  observed  in  persons  handling 
these  parasites;  aldehydes  are  notoriously^  inclined  to  produce  con- 
ditions of  hypersensitiveness,  e.  g.,  formaldehyde.  It  is  quite  possi- 
ble that  the  severe  constitutional  symptoms  observed  occasionall}^  in 
persons  infected  with  ascaris,  are  produced  by  these  substances  or 
by  poisonous  substances  set  free  through  disintegration  of  worms 
which  have  died  and  remained  in  the  bowel.  A  capillary  poison  re- 
sembling sepsin,  poisonous  bases  acting  like  atropine  and  coniine, 
and  hemolytic  unsaturated  fatty  acids  were  also  found,  among  other 
less  toxic  substances  produced  by  ascaris,  and  the  sum  of  their  ac- 
tion is  certainly  adequate  to  account  for  anything  ascribed  to  these 
parasites.  Paulian,^^'^  however,  would  attribute  the  chief  effect  to 
anaphylaxis  from  absorbed  proteins,  while  Brincla  '''^  believes  that 
ascaris  produces  an  active  toxalbumin.  This,  he  found,  causes  a 
tetany-like  type  of  respiration,  and  a  similar  symptom  is  often  noticed 
in  children  with  ascarides.  An  actively  toxic  mixture  of  proteoses  and 
peptones  has  been  obtained  from  several  species  of  ascaris,  and  desig- 
nated as  "askaron, "  by  Shimamura  and  Fujii."'' 

Analysis  of  a  great  quantity  of  ascaris  from  liorse  and  liog  gave  as  tlie  chief 
results,  the  following:  ^7  They  differ  much  in  composition  from  the  higher  ani- 
mals. About  half  the  ash  is  water  soluble;  and  of  tlie  dry  substance  about  half 
is  protein  or  related  substances,  from  which  the  usual  amino-acids  and  purines 
can  be  isolated.  Uric  acid  and  creatinin  were  lacking.  The  superficial  layer  does 
not  consist  of  chitin,  but  of  an  albuminoid  rich  in  sulphur  and  free  from  carbohy- 
drates, resembling  keratin.  They  have  abundant  and  active  enzymes  of  many 
kinds.  Glycogen  is  the  chief  carbohydrate,  but  there  are  also  glucoproteins  and 
gluc^se.  The  ascaris  differs  from  higher  animals  especially  in  the  ether-soluble 
substances,  which  consist  chiefly  of  free  fatty  acids,  many  of  which  arc  volatile. 
Also  found  were  lecithin,  aldehydes  and  neutral  fats,  but"  little  glycerol,  no  chol- 
esterol, and  an  '"ascaryl  alcohol''  (C.oHo,  0.)  which  probably  substitutes  for  both 
glycerol  and  cholesterol. 

Trichinella  Spiralis  has  been  investigated  from  the  chemical  stand- 
point by  Flury,''*  who  found  that  the  infected  nniseles  of  experimental 

57  Arch.  exp.  Path.  u.  Pharm..  ini2    (G7K  275    (literature). 
57aCompt.  Rend.  Soc.  Biol.,   1915    (78),  73. 

57b  Arch,  de  M6d.,   1915    (17),  SOI. 

57c  Japanese  Jour.  Pact.    (Saikingaku  Zassi),  June  10.   1016. 

58  Arch.  exp.  Path.  u.  Pharm.,  1913    (73),  164  and  214. 


142  CHEMISTRY    OF    THE    ANIMAL    PARASITES 

animals  differed  from  normal  muscles  in  having  more  water  because 
of  edema,  an  increase  in  extractives,  ammonia  coinpounds,  lactic  acid 
and  volatile  acids,  with  fluctuating  values  in  both  creatine  and  purines, 
(xlyeogen  is  decreased  not  only  in  the  infected  muscle  but  also  in  the 
liver  and  kidneys.  The  parasites  themselves  are  remarkably  resistant 
to  strong  acids,  perhaps  because  of  the  lipoid  content  of  their  surface 
covering,  in  which  keratin  could  not  be  positively  identified;  choles- 
terol and  glycogen  were  present.  The  blood  of  infected  animals  shows 
an  excess  of  nuclein  material,  and  may  give  albumose  and  diazo  re- 
actions; the  red  corpuscles  have  a  lowered  resistance  to  hemolysis  by 
hypotonic  solutions.  Trichinous  muscle  contains  substances  that  pro- 
duce marked  local  tissue  irritation,  which  may  be  purines;  a  curare- 
like poison  was  also  found,  which  was  believed  to  be  a  guanidine  deriva- 
tive, as  well  as  a  "fatigue  poison"  which  probably  consists  of  the 
lactic  acid  and  other  muscle  extractives.  The  location  of  trichinella 
in  muscle  may  be  ascribable  to  their  need  for  glycogen  for  nourish- 
ment and  the  fact  that  their  metabolism  is  carried  out  anaerobically 
may  account  for  the  character  of  the  products  (fatty  acids,  etc.). 

The  intoxication  of  trichinosis  probably  is  the  combined  result  of 
the  products  of  the  metabolism  of  the  parasites,  the  products  of  muscle 
disintegration,  and  perhaps  also  of  anaphylactic  reaction  to  the  pro- 
teins of  the  parasite  and  the  altered  muscle  proteins.  As  evidence  of 
the  anaphylactic  condition  is  the  conspicuous  eosinophilia,  which  we 
know  is  often  the  result  of  anaphylactic  intoxication,-'**''  Metabolism 
studies  show  a  preliminary  nitrogen,  creatinine  and  purine  retention, 
followed  by  excessive  loss  of  all  three.  There  is  also  an  intense  diazo 
reaction,  and  increased  excretion  of  lactic  and  organic  acids.  The 
hypothesis  that  bacterial  invasion  is  responsible  for  the  intoxication 
of  trichinosis  does  not  seem  to  be  well  supported  (Herrick,  Gruber). 

The  serum  of  infected  animals  is  not  toxic,  and  does  not  protect 
against  infection  with  trichinella  (Gruber^*'').  Salzer,^*''  however, 
found  that  the  serum  of  recovered  patients  had  a  curative  effect  in 
persons  acutely  intoxicated  with  trichiniasis,  and  also  a  marked  pro- 
phylactic effect  in  experimental  animals;  it  removed  the  eosinophilia 
both  in  men  and  animals.  lie  also  observed  evidence  of  a  reduction 
of  the  bilirubin  of  the  feces  by  the  trichinae,  so  that  the  stools  were 
clay  colored  without  icterus.  Positive  complement  fixation  reactions 
are  given  by  tlio  serum  of  trichinella  infected  poreons. '*''''' 

TJncinaria  duodenalis,  wliieh  has  for  its  chief  effect  tlie  production 
of  a  severe  anemia,  seems  to  cause  this  anemia  by  producing  rejieated 
small  hemorrliages  rather  than  by  any  toxic  action.     Tlie  abundance 

5Ra  Spo  llcrrick,  Jour.  Amor.  ^Icd.  Assoc,  ini,5  ((>")),  ISTfl;  Schwartz.  Kri;i'b. 
allp.  Pat.liol.,  lit  14   (17),  l.^ti. 

BSbMiincli.  med.  Wocli.,  1!)14    (01),  045. 
58r.Tour.  Amer.  Mod.  Assoc,  1010   (67),  .')79. 
58dStroebel,  Miinch.  mod.  Woch.,   1911    (58),  G72. 


NEMATODES  143 

of  tliis  loss  of  blood  is  explained  by  L.  Loeb  '''''  as  due  to  the  presence, 
in  the  anterior  portion  of  the  parasite  (they  studied  Ankylostonia 
caniniim),  of  a  substance  that  inhibits  the  coa^lation  of  the  blood. 

However,  Preti  "^  would  ascribe  importance  to  a  lipoidal  or  lipoid- 
like  hemolytic  constituent  of  the  parasitic  tissues  of  the  European 
ankylostoma,  but  Whipijle,*"'^  who  lias  observed  a  weak  hemolysin  in 
the  American  hook  worm,  considers  it  too  ineffective  to  be  of  practical 
importance.  In  Sclerostoma  equinmn,  however,  Bondonoy  "-  found 
active  hemolytic  agents,  ascribed  by  him  to  lipase ;  also  a  ptomain,  an 
alkaloid  and  other  substances.  Correspondinfj  to  Flury's  analyses 
of  ascaris,  he  found  that  the  cuticle  is  albuminoid  and  not  chitinous, 
and  that  the  parasite  produces  much  volatile  fatty  acids,  especially 
butyric;  both  lecithin  and  cholesterol  were  absent.  The  dermatitis 
produced  by  uncinaria  larva?  is  ascribed  by  C.  A.  Smith  ^^  to  an  alco- 
hol-soluble substance.  Wateiy  extracts  of  Sclerostoma  were  found  by 
Grosso  "*  to  cause  but  slight  chemotaxis  without  eosinophilia. 

Filaria  seem  not  to  produce  any  appreciable  amount  of  toxic  ma- 
terial, if  we  may  judge  by  the  slight  evidence  of  intoxication  shown 
by  infected  individuals.  An  exception  may  be  made  in  the  case  of 
the  guinea-worm  {Dracunculus  or  F.  medinensis) .  This  parasite 
causes  chiefly  mechanical  injury  unless  its  body  is  ruptured,  which 
may  happen  in  attempting  to  remove  it  forcibly;  this  accident  is  fol- 
lowed by  violent  local  inflammation  or  gangrene,  which  indicates 
that  some  powerfully  irritant  substance  is  liberated  from  the  torn 
body  of  the  worm.^^ 

59  Cent.  f.  Bakt.,  1004  (37),  93;  1906  (40),  740;  Loeb  and  Fleischer,  Jour. 
Infec.  Dis.,  1910    (7),  625. 

eoMiinch.  med.  Woch.,  1908    (5.5).  436. 

61  Jour.  Exp.  Med.,  1909   (11),  331. 

62  Arch.  Parasitol.,  1910  (14),  5:  see  also  Ashcroft,  Compt.  Rend.  Soc.  Biol., 
1914   (77),  442. 

63  Jour.  Amer.  Med.  Assoc,  1906    (47),   1693. 

64  Folia  Hematol.,  1912    (14),  18. 

65  Earthworms  are  said  by  Yagi  (Arch,  internat.  phaimacodyn..  1911  (21), 
105)  to  contain  a  hemolytic  substance,  "lumbricin,"  the  properties  of  -which  he 
describes.  Nukada  and  Tenaka  (Mitt.  med.  Fakult.,  Tokio.  1915  (14),  1),  found 
an  antipyretic  agent  which  seems  to  be  derived  from  tyrosine. 


CHAPTER    VI 

PHYTOTOXINS  AND  ZOOTOXINS 

The  production  of  substances  possessing  the  essential  features  of 
true  toxins  is  by  no  means  limited  to  the  bacterial  cell.  In  the  plant 
kingdom  such  substances  are  formed,  and  called  phytotoxins.  Of 
these,  the  best  known  are  ricin,  abrin,  crotin,  and  robin.  Among 
the  toxins  of  animal  origin,  zootoxins,  are  the  venoms  of  poisonous 
snakes,  lizards,  spiders  and  scorpions,  and  the  serum  of  eels  and 
snakes.     These  may  be  briefly  considered  as  follows : 

PHYTOTOXINS 

The  chief  phytotoxins  ^  are  as  follows : 
Ricin,  from  the  castor-oil  bean  {Bicinus  communis) . 
Abrin,  from  the  seeds  of  Ahrus  prccatorius. 
Crotin,  from  the  seeds  of  C  rot  on  tigliiim. 

Robin,  from  the  leaves  and  bark  of  the  locust,  Bolinia  pseudo- 
acacia. 
Curcin,  from  the  seeds  of  Jatropha  curcus. 

In  their  general  properties  all  these  substances  are  very  similar  and 
may  be  considered  together.  They  resemble  proteins  in  many  re- 
spects, for  they  can  be  salted  out  of  sohitions  in  definite  fractions 
of  the  precipitate,  are,  precipitated  by  alcohol,  and  are  slowly  de- 
stroyed by  proteolytic  enzymes.  For  some  time  they  were  referred 
to  in  the  literature  as  toxalbumins,  until  Jacoby  stated  that,  by  com- 
bining the  salting-out  method  with  trypsin  digestion,  he  was  able 
to  secure  preparations  of  ricin  and  abrin  that  did  not  give  the  pro- 
tein reactions.  This  seemed  to  pUice  tliem  in  the  same  category 
with  bacterial  toxins  and  enz.ymes,  /.  r.,  large  molecular  colloids,  closely 
resembling  the  proteins  with  which  they  are  associated,  but  still  not 
giving  the  usual  protein  reactions.  Because  of  their  great  similarity 
to  bacterial  toxins  tliis  seemed  a  very  probable  description,  and  it  has 
been  generally  accepted.  More  recent  work  by  Osborne,  Mendel,  and 
Harris,-  however,  does  not  support  Jacoby 's  contention.  They  found 
the  toxic  properties  of  ricin  associated  inseparably  with  the  coagulable 
albumin  of  the  castor  beans,  and  were  able  to  isohite  it  in  such  purity 

1  Rr-suni(5  of  litpi-atuiv  hv  Ford,  Cent.  f.  Bakt.,  101.3  (aS).  12!);  .lacohv.  Kollo 
and  Wasscrmann's  Iland))uch.   1!)1,3    (2),    14;-).'?. 

2  Amcr.  Jour,  of  Plivsiol.,   100.')    (14).  2.'i0. 

144 


PHYTOTOXIN  IMMIXITY  145 

that  one  one-thousandth  of  a  milligram  (0.000001  gram)  was  fatal  per 
lilo  of  rabbit,  and  solutions  of  0.001  per  eent.  would  agglutinate  red 
corpuscles.  The  toxicity  was  also  impaired  or  destroyed  by  tryptic 
digestion.  They  consider  that  probably,  because  of  its  extremely 
great  toxicity,  Jacoby  was  able  to  get  active  preparations  that  con- 
tained too  little  active  substance  to  give  the  protein  reactions.  As 
they  remark:  "If  one-thousandth  of  a  milligram  of  a  compound 
giving  on  analysis  every  indication  of  being  a  relatively  pure  protein, 
is  phj^siologically  active  in  the  degree  characterized  by  our  experiments, 
the  toxicity  of  any  impurity  must  be  infinitely  greater  than  tliat  of 
any  known  toxins."  Against  the  claim  that  the  toxic  princii)le  is 
simply  carried  down  with  the  protein  is  the  fact  that  it  does  not  come 
down  in  the  first  fraction  that  is  precipitated,  the  globulin,  which  usu- 
ally carries  down  all  impurities.  All  the  ricin  comes  down  between 
the  limits  of  one-fifth  and  one-third  saturation  with  ammonium  sul- 
]ihate,  exactly  as  does  the  albumin.  Field  ^  has  found  evidence  that 
the  agglutinin  and  toxin  of  pure  ricin  are  separable,  but  Reid  believes 
them  identical.  Of  21  varieties  of  ricinus  seeds  examined  by  Agul- 
hon,^^  all  yielded  hemagglutinins.  During  germination  of  the  castor 
bean  the  ricin  disappears  with  the  albumin.^''  Ricin  agglutinates  not 
only  corpuscles,  but  tissue  cells  of  all  sorts,  and  causes  precipitates  in 
normal  serum.'*  Curcin  alone  seems  to  have  no  hemagglutinative 
action.*^ 

Immunity. — The  phytotoxins  have  been  very  serviceable  in  the 
study  of  immunity,  since  they  obey  the  same  laws  as  bacterial  toxins 
and  can  be  handled  in  more  definite  quantities.  By  their  use  Ehrlich 
first  determined  that  toxin  and  antitoxin  act  quantitatively.  They 
seem  to  possess  haptophore  and  toxophore  groups,  and  immunity  is 
readily  obtained  against  them,  not  only  by  subcutaneous  injection, 
but  by  dropping  into  the  conjunctival  sac,  and  also  by  feeding,  show- 
ing their  direct  absorbability  and  their  resistance  to  digestion.  The 
antitoxin  is  present  in  the  milk  of  the  immunized  mother  and  im- 
munizes the  suckling;  but  little  is  carried  through  the  placenta  into 
the  fetal  blood.  The  immunity  is  specific,  ricin  antitoxin,  for  exam- 
ple, not  protecting  against  abrin  (although  it  is  said  to  protect  against 
robin).  Roemer  found  that  in  animals  immunized  by  conjunctival 
application  the  eye  so  used  became  immune  to  the  local  action  of  the 
poison  before  the  other  eye  did,  indicating  a  local  development  of 
immune  substance.  In  general  immunization  the  immune  substance 
appears  first  in  the  spleen  and  bone-marrow.  Normal  serum  gives 
a  precipitate  with  ricin,  but  immune  serum  gives  a  much  heavier  one. 

3  Jour.  Exper.  Med.,  1910  (12),  o.il ;  Keid,  Landwirtsch.  Versuchst..  101.3  (F2), 
393. 

3a  Ann.  Inst.  Pasteur.  1014   (28).  SIO. 
3bA<TuIhon,  ibid.,  101.5    (20).  2.37. 

4Michaelis  and  Steindorff.  Biocliem.  Zeit..  1000    (2),  43. 
4aFelke,  Landwirts.  Versuchst.,  1913    (82),  427. 
10 


146  PHYTOTOXINS    AND    ZOOTOXINS 

Antiricin,  like  other  antitoxins,  is  inseparable  from  the  proteins  of  the 
serum. 

Physiological  Action. — Their  poisonous  action  is  manifold,  most 
prominent  being-  aggiutination  of  the  erj'throcytes,  local  cellular  de- 
struction, and,  to  a  less  extent,  hemolysis.  Jacoby  believes  that  in 
ricin  there  are  several  toxic  substances  differing  in  physiological  prop- 
erties, similar  to  Ehrlich's  findings  in  diphtheria  toxin  (toxones, 
etc.).  By  long  action  of  pepsin-HCl  upon  ricin,  he  secured  a  prepa- 
ration with  all  the  other  properties  of  ricin  except  that  it  was  inactive 
against  erythrocytes ;  the  same  result  could  not  be  obtained  with 
abrin.  Heating  to  65°  or  70°  does  not  destroy  the  toxicity  of  phy- 
totoxins,  but  boiling  does.  There  is  a  latent  period  of  several  hours 
after  injection  of  the  poison,  the  onset  of  symptoms  being  sudden; 
death  rarely  occurs  in  less  than  fifteen  to  eighteen  hours  (Osborne 
et  al.). 

Flexner  ^  has  studied  particularly  the  histological  changes  pro- 
duced by  ricin  and  abrin  poisoning  in  animals.  Both  act  alike,  af- 
fecting the  tissues  much  as  bacterial  toxins  do  (diphtheria).  Fever, 
albuminuria,  and  convulsions  are  followed  by  exhaustion  and  lowered 
temperature.  Punctiform  hemorrhages  are  found  beneath  the  serous 
surfaces,  with  fluid  in  the  peritoneal  cavity.  At  least  in  the  case 
of  ricin  the  hemorrhages  are  not  due  to  blood  changes,  but  to  a  spe- 
cial toxin  destroying  the  endothelial  cells."  There  occur  a  general 
lymphatic  enlargement  and  marked  changes  in  the  intestinal  mucosa, 
Avith  swelling  of  the  Beyer's  patches.  The  spleen  is  swollen  and 
dark  in  color,  as  also  is  the  liver,  which  shows  much  focal  necrosis. 
The  glycogen  content  of  the  liver  is  decreased  in  abrin  poisoning.'' 
Subcutaneous  injection  causes  local  edematous  inflammation  without 
suppuration.  Histologically,  in  the  most  affected  organs  are  found 
much  cellular  necrosis  and  disintegration,  especially  of  lymphoid  and 
epithelial  cells.  Changes  in  the  capillary  endothelium,  fibrinous 
thrombi,  and  abundant  hemorrhagic  extravasations  are  widespread. 
Probably  agglutinative  thrombosis  by  red  corpuscles  plays  an  im- 
portant part  in  these  intoxications  (Ehrlich),  but  Aschoff ''^  ascribes 
the  thrombosis  to  the  fragments  of  disintegrated  marrow  and  blood 
cells.  The  great  amount  of  intestinal  injury  probably  depends  upon 
the  fact  that  these  poisons  are  largely  eliminated  through  the  intestinal 
inucosa.  There  are  also  severe  changes  in  the  bone  marrow,  accom- 
panied by  the   appearance  of  nucleated  erythrocytes  in  the  blood.* 

Mushroom  Poisons. — -Tlio  poisons  of  tlie  three  cliiof  poisonous  iimslirooms. 
Arnanita  miiscaria,  Ilelvella  esciilevtia,  and  Amanita  phalloidrs,  dift'er  from  one 
anotlier    quite    essentially.     The    poisonous    principles    of    the    first    and    second, 

-Jour.  Exper.  Med.,  1897    (2),  197. 

6  Amer.  Jour.  Med.  Sci.,  1903    (126).  gOG. 

T  Dovon,  Conipt.  Rend.  Soc.  Biol.,  1909    (07).  .SO. 

TaArch.  Int.  Med..   1913    (12),  503. 

^  Buntin},',  Jour.  Kxper.  :Mcd.,  1900    (S),  025. 


HAT  FEVER  147 

muscarine  and  lielvellic  acid,  are  non-protein  substances,  of  known  cliemical  com- 
position, wliich  are  discussed  elsewlierc;  but  the  Amanita  phailoidcn,  tlie  most 
important  of  the  tiiree,  owes  its  toxic  properties  to  at  least  two  poisonous  con- 
stituents. One  is  powerfully  hemolytic,  is  destroyed  by  heatinff  tliirty  minutes  at 
65°,  and  acts  directly  upon  red  corpuscles  without  the  presence  of  s(!runi.'^" 

The  studies  of  Ford'-'  aiul  his  associates  have  shown  that  this  hemolysin  is  a 
glucoside,  yielding  on  hydrolysis  pentose  and  volatile  bases,  and  yet  capable  of 
acting  as  an  antigen,  since  actively  antihemolytic  sera  can  be  produced  by  im- 
munizing animals.  This  substance  corresponds  to  the  phallin  of  Koljcrt. 
Probably  this  hemolytic  poison  is  not  the  important  agent  in  poisoning 
by  Amanita,  as  it  is  easily  destroyed  by  heat  and  the  digestive  fluids.  The  ther- 
mostable poison,  Amanita- toxin,  gives  no  reactions  for  either  glucosides  or  pro- 
teins,^o  and  does  not  confer  any  considerable  antitoxic  property  on  the  blood  of 
immunized  animals.  The  toxin  kills  acutely,  the  animals  dying  in  24-48  hours, 
and  show'ing  no  changes  beyond  a  fatty  degeneration  of  the  internal  organs.  Tlie 
hemolysin  kills  slowly  in  .VI 0  days,  causing  local  edema  aiul  liemoglobinuria. 

Amanita  muscaria  contains  a  heat-resistant  agglutinin  which  also  seems  to  be  a 
glucoside,  but  it  is  not  toxic  nor  antigenic. 

An  extensive  study  of  many  fungi  by  Ford  n  led  him  to  classify  the  toxic  action 
in  three  groups:  (1)  nerve  poisons,  e.  g.,  muscarine;  (2)  those  causing  struc- 
tural changes  in  the  viscera,  e.  g.,  A.  phalloides.  causing  fatty  degeneration;  (3) 
gastro-intestinal  irritants,  e.  g.,  Lactarius  torminosits. 

The  poison  of  Rhus  toxicodendron  has  also  been  found  by  Aeree  and  Syme  12  to 
be  a  glucoside,i2a  and  the  same  is  true  of  the  poison  oak,  Rhus  diversiloha,  which 
has  no  antigenic  properties. is 

HAY-FEVER 

In  1902  Dunbar  i*  demonstrated  conclusively  that  typical  hay-fever,  in  its 
several  various  forms,  is  due  to  pollen  of  various  sources,  in  all,  twenty-five 
varieties  of  grass  and  seven  varieties  of  plants  of  other  sorts  being  found  whose 
pollen,  when  placed  upon  the  nasal  or  conjunctival  mucous  membranes  of  hay- 
fever  patients,  causes  a  typical  attack  of  the  disease.  In  Germany  the  disease 
seems  to  come  chiefly  from  pollen  of  the  grasses  and  grains  (rye  pollen  being 
most  active),  whereas  in  America  the  most  important  pollen  seems  to  come  from 
members  of  the  Ambrosia  (rag-weed)  and  SoUdago  (goldenrod)  .i*a  Dunbar  also 
found  tliat  tlie  toxic  constituent  could  be  dissolved  from  the  pollen  in  salt  solu- 
tion, and  seemed  to  be  a  protein.  The  protein  constituents  of  the  pollen  of  rye 
have  been  studied  further  by  Kammann,io  who  found  three  proteins,  one  of  which, 
an  albumin,  was  found  to  contain  all  the  active  matter.  This  constitutes  about 
5.5  per  cent,  of  tlie  entire  weight  of  the  pollen,  is  weakened  but  little  by  heating 
to  80'°,  and  is  not  destroyed  bv  boiling;   it  is  but  partly  destroyed  by  pepsin  and 

1 

trypsin,  and  resists  acids  but  not  alkalies.     A  solution  containing   • milligram 

120 
of  pollen  protein,  which  amount  is  contained  in  two  or  three  pollen  grains,  pro- 
duces a  reaction  in  susceptible  individuals,  but  large  amovmts  have  no  effect  on 
normal  persons.  Dunbar  has  manufactured  an  antitoxic  seriun  by  immunizing 
horses  against  the  pollen,  although  by  no  means  all  observers  are  agi-eed  as  to 
its  efficacy. 

8a  The  hemagglutinin  of  Agaricus  campestris  is  precipitated  at  a  H-ion  concen- 
tration of  2.6  X  10-*   (Brossa,  Arch.  sci.  med.,  1915   (39),  241). 
9  See  Jour,  of  Pharm.,  1910   (2),  145;   1913   (4),  235,  241  and  321. 
loRabe   (Zeit.  exp.  Path.,  1911    (9),  352)   considers  it  to  be  an  alkaloid. 

11  Jour,  of  Pharm.,  1911    (2),  285. 

12  Jour.  Biol.  Chem.,  1907    (2),  547. 

12a  Questioned  by  McNair,  Jour.  Amer.  Chem.  Soc,  1916   (38),  1417. 

13  Adelung,  Arch.  Int.  Med.,   1913    (11),   148. 

14  Full  review  of  subject  and  literature  given  by  Prausnitz,  KoUe  and  Wasser- 
mann's  Handbuch,  1913  (2),  1469;  Koessler,  Forchheimer's  Therapeutics,  1914 
(5),  671. 

14a  See  Cooke  and  Van  der  Veer,  Jour.  Immunol.,  1916  (1),  201;  Goodale,  Bos- 
ton Med.  Surg.  Jour.,  1914   (171),  695. 

15  Hofmeister's  Beitr.,  1904   (5),  346;  Biochem.  Zeit.,  1912   (46),  151. 


148  PHYTOTOXINS    A^D    ZOOTOXINS 

The  processes  involved  in  hay  fever  would  seem  to  be  characteristically  in  the 
nature  of  an  anaphylactic  reaction  to  a  foreign  protein,  in  which  case  we  cannot 
speak  of  either  a  toxin  or  an  antitoxic  seriun.  Prausnitz  suggests  that  the  serum 
of  immunized  animals  contains  so  large  a  supply  of  antibodies  that  the  minute 
quantities  of  foreign  protein  are  digested  beyond  the  toxic-fraction  stage  almost 
immediately,  and  hence  no  reaction  is  observed.  This  explanation  would  agree 
with  the  experimental  inhibition  of  the  toxicity  of  pollen  mixed  witli  antiserum, 
and  at  the  same  time  with  the  clinical  inefficiency  of  the  serum.  The  hyper- 
sensitization  seems  to  be  established  spontaneously  through  inheritance,  but  no 
antibodies  can  be  demonstrated  in  the  blood  of  sensitized  persons,  altliougli  the 
cells  of  the  skin  and  mucous  membranes  are  reactive.io 

(The  effects  of  the  phytotoxins  on  the  blood  are  discussed  under 
"Hemolysis"  in  Chapter  viii.  Vegetable  hemolytic  poisons  that  do 
not  resemble  the  toxins,  e.  g.,  giucosides,  etc.,  will  also  be  found  dis- 
cussed under  the  same  heading.) 

ZOOTOXINS  18 

SNAKE  VENOMS  19 
This  important  class  of  poisons,  first  thoroughly  investigated  by 
AVeir  Mitchell  (1860),  and  :\Iitchell  and  Reichert  (1883),  has  re- 
cently aroused  great  interest  through  its  relations  to  bacterial  toxins 
and  the  problems  of  immunity.  The  poisons  of  different  species  of 
snakes  seem  to  have  much  in  common  with  one  another,  whether  de- 
rived from  the  Elaperine  snakes  (cobras  and  numerous  other  Indian 
and  Australian  snakes),  or  Viperidce  (including  most  poisonous  Amer- 
ican snakes),  or  Hydrophime  (the  poisonous  sea-snakes),  although 
very  characteristic  differences  exist  between  each. 

The  essential  anatomical  differences  between  the  different  classes  of  snakes  are 
as  follows:  Colubrid<c,  which  include  all  the  non-poisonous  snakes,  have  no 
mechanism  for  injecting  poisons  into  their  victims.  Cohihrid<i'  renenosce  are 
venomous  snakes  resemi)ling  in  many  particulars  the  harmless  Colubrines,  but 
having  short  poison  fangs,  firmly  fastened  to  the  maxilla  in  an  erect  position; 
in  this  class  are  included  the  cobra  and  the  venomous  snakes  of  Australia. 
Viperidce,  or  vipers,  are  characterized  by  a  highly  specialized  apparatus  for  in- 
jecting the  poison;  their  poison  fangs  are  very  long,  and  the  miixillary  bone,  to 
which  they  are  fastened,  is  so  articulated  that  it  rotates  about  a  (|u;n-ter  of  a 
circle  when  the  snake  strikes,  bringing  the  fangs  into  an  erect  position.  Tlie 
fangs  are  canalized  and  pointed  at  the  end  like  a  hypodermic  needle,  and  the 
poison  is  forced  through  them  under  considerable  pressure  by  a  large  muscle  that 
contracts  over  the  salivary  gland.  Accessory  fangs  in  various  stages  of  develop- 
ment are  also  present  to  replace  any  fang  lost  in  action.  All  the  poisonous 
snakes  of  North  America,  with  one  insignificant  exception,  belong  to  the  vipers, 
and  to  a  special  class  known  as  the  "pit  vipers,"  because  of  the  presence  of  a 
deep  pit  of  unknown  function  above  the  maxilla.  The  exception  mentioned  is 
the  "coral  snake"  found  on  the  coast  of  Florida,  around  the  Gulf  of  :Mexico  and 

10  See  Cooke,  Flood  and  Coca,  Jour.  Immunol.,  1917    (2),  217. 

18  Full  review  and  literature  given  by  Faust,  "Die  tierischen  Gifte,"  Braun- 
schweig, 1906;  also  in  Abdorhalden's  Ilandbuch,  Vol.  II.  Sachs,  Kolle  and  Was- 
sermann's  Handbuch,  1913    (2),  1407. 

19  Elaborate  review  and  bibliography  given  by  Noguchi,  Carnegie  Institution 
Publications,  1900,  No.  Ill:  also  bv  Calmette,  "Lea  v<^nins.  les  animaux  veriimaux 
et  la  s^-rotherapie  anlivenijueusc."  Paris.  IMasson,  1907:  Calmette,  Kolle  ami  Was- 
sermann's  Handbuch,  Vol.  IT,  j).  1.S81;  witli  reference  1o  Xortli  American  snakes, 
see  Prentiss  Willson,  Arch.  Int.  :\red.,  1908   (1),  516. 


SNAKE  VENOMS  149 

in  the  southeastern  states;  it  is  a  member  of  the  coliibrine  poisonous  snakes,  of 
small  size,  and  seldom  causes  serious  poisoning.  The  poisonous  vipers  are  the 
rattlesnakes  (Crotaliis) ,  of  which  there  are  some  ten  to  twelve  or  more  species, 
and  Sistrtirus,  of  whicli  there  are  two  species:  the  copperheaded  adder  (Ancistro- 
don  contortrix)  and  the  water  moccasin  {Ancistrodon  piscivorus) . 

The  classification  used  above  is  the  one  followed  in  most  publications  on 
poisonous  snakes;  a  more  modern  classification  divides  the  snakes  (Ophidia)  into 
several  series,  one  of  these  includino;  all  poisonous  snakes  under  the  title  of 
Proterogiypha,  and  dividing  this  series  into  the  three  families:  (1)  Elapinw, 
including  cobras,  coral  snakes,  etc.;  (2)  Ilydrophinije,  the  poisonous  sea-snakes; 
(3)    Viperidic,  including  all  snakes  with  erectile  fangs. 20 

The  source  of  the  venom  is  probably  in  part  the  blood,  since  snake 
blood  has  been  found  to  contain  poisons  very  similar  to  some  of  those 
in  the  venom ;  therefore  these  are  presumably  simply  filtered  out 
by  the  venom  glands,  and  not  manufactured  by  them.-^  Other  poison- 
ous constituents  of  venom  are  not  found  in  snake  serum,  and  there- 
fore are  probably  manufactured  by  the  venom  gland.  Apparently 
many  of  the  harmless  snakes  produce  a  poisonous  saliva,  since 
extracts  of  their  glands  are  said  hj  Blanchard  --  to  possess  the 
properties  of  the  venoms,  and  if  so  these  snakes  are  harmless  chiefly 
because  they  lack  an  apparatus  for  injecting  the  poison.  As  a  rule, 
however,  the  venom  glands  are  much  more  highly  developed  in  the 
poisonous  snakes,  and  are  connected  with  a  specialized  injection  ap- 
paratus :  in  structure  they  are  compound  racemose  glands. 

Properties  of  Venom. — ^As  ejected,  the  venom  is  weakly  acid  or 
neutral  in  reaction,  and  free  from  bacteria,  contrary  to  earlier  ideas 
(Langmann).  Its  specific  gravity  is  1030  to  1077,  and  it  contains  a 
large  amount  of  solids,  generally  20  to  40  per  cent,  by  weight.  These 
are  precipitated  by  alcohol,  ether,  tannin,  and  iodin,  but  do  not  ad- 
here to  precipitates  of  phosphates  as  do  enzymes  and  toxins  (Cal- 
mette).  They  do  not  diffuse  through  dialyzing  membranes.  When 
dried,  the  venom  can  be  kept  almost  indefinitely  without  losing  its 
strength,  specimens  over  twenty  years  old  having  been  found  unim- 
paired. Glycerol  and  alcohol  also  seem  not  to  injure  it,  but  oxidiz- 
ing agents  of  all  kinds  are  very  destructive.  Light  impairs  the  power 
of  venoms,  as  also  does  radium  (Phisalix).-^  Eosin  and  erythrosin 
also  reduce  the  power  of  venom  through  their  photodynamic  action, 
affecting  the  neurotoxic  properties  less  than  the  hematotoxic  compo- 
nents (Noguchi).^*  Cobra  venom  withstands  even  100°  for  a  short 
time,  but  crotaline  venoms  are  destroyed  at  80-85°. 

20  For  a  full  discussion  of  the  characteristics  of  the  poisonous  snakes  of  North 
America,  see  the  monograph  with  that  title  by  Stejneger,  Report  of  U.  S.  Na- 
tional jNIuseum,  189.3,  Washington.  A  good  summary  is  also  given  by  Langmann, 
Reference  Handbook  of  INIedical  Sciences.  Concerning  poisonous  sea-snakes, 
Ilydrophidia,  see  Boulanger,  Natural  Science,  1802  ( 1) .  44.  The  poisonous  snakes 
of  India  are  described  by  Fayrer,  in  "The  Tlianatopliidia  of  India."'  London,  1874. 

21  Contradicted  by  Arthus,  Arch,  internat.  phvsiol.,  1012   (12),  1G2. 
22Compt.  Rend.  Soc.  Riol.,  1894    (46),  3.5. 

23  Compt.  Rend.  Soc.  Biol.,  1904    (.56),  327. 
24, Jour.  Exper.  Med.,  1906   (8),  252. 


150  PEYTOTOxiyn  axd  zootoxins 

Much  work  has  been  done  upon  the  nature  of  the  constituents  of 
venom.  As  early  as  1843  Prince  Lucien  Bonaparte  found  that  there 
were  proteins  in  the  venom,  which  was  corroborated  by  ]\Iitchell  in 
1861.  In  1888  Mitchell  and  Reichert  described  two  poisonous  pro- 
tein constituents  of  venom,  one  of  wliich  was  eoagulable  by  heat  and 
seemed  to  be  a  globulin;  the  other  resembled  the  proteoses  (they 
called  it  "peptone,"  according-  to  the  nomenclature  of  that  time). 
To  the  globulin  they  ascribed  the  local,  irritating  properties  of  venom ; 
to  the  albumose,  the  systemic  intoxication.  Corresponding  to  their 
action,  venoms  of  different  serpents  were  found  to  vary  greatly  in  the 
proportions  of  these  proteins.  Cobra  venom,  which  acts  chiefly  sys- 
temieally,  contains  98  per  cent,  of  albumose  and  but  2  per  cent,  of 
globulin;  rattlesnake  venom,  with  its  marked  local  effects,  contains 
25  per  cent,  of  the  irritating  globulin ;  moccasin  venom  contains  8 
per  cent,  of  globulin.  Several  other  observers  soon  corroborated  the 
main  facts  of  Mitchell  and  Reichert 's  report;  but,  as  has  been  seen  in 
connection  with  the  consideration  of  the  composition  of  enzymes,  tox- 
ins, etc.,  the  fact  that  a  substance  is  carried  down  with  a  protein  is 
no  proof  that  it  is  itself  a  protein.  What  has  been  established  is 
merely  that  the  irritating  component  of  venom  can  be  destroyed  by 
heat,  and  is  removed  with  the  globulin  in  fractional  separation ;  while 
there  remains  a  substance  not  destroyed  by  boiling,  which  comes  down 
at  least  in  part  with  the  albumoses  of  the  venom,  and  causes  chiefly 
systemic  manifestations. 

Since  venoms  act  as  antigens  and  stimulate  the  formation  of  spe- 
cific antibodies,  it  is  to  be  presumed  that  the  poisonous  principles 
are  proteins,  or  toxalbumins,  although  this  conclusion  does  not  neces- 
sarily follow.  Faust  -^  believes  the  poison  of  venoms  not  to  be  pro- 
teins, but  glucosides,  free  from  nitrogen,  resembling  very  much  quil- 
lajic  acid,  and  therefore  belonging  to  the  saponin  group  of  hemolytic 
agents.  He  has  isolated  such  a  substance  from  cobra  venom,  which 
he  calls  ophiotoxin  (C^-HooOio),  and  from  rattlesnake  venom  a  sub- 
stance which  seems  to  be  a  polymer  of  the  ophiotoxin,  (C.54H-4O21). 
Possibly  these  glucosides  are  bound  to  proteins,  forming  compound 
proteins  which  act  as  specific  antigens.  According  to  this  work  the 
snake  venoms  and  the  dermal  poisons  of  toads  and  frogs  are  all  closely 
related  substances. 

Enzymes  in  Venoms. — As  venom  causes  raiiid  liquefaction  of  tissues 
into  wliich  it  is  injected,  Flexner  and  Nognclii  -"  tested  crotalus  and 
cobra  venom  for  proteases,  and  found  that  tliey  digested  nuiscle  rap- 
idly, and  also  gelatin  and  unboiled  fibrin ;  whereas  boiled  fibrin  and 
boiled  egg-albumen  were  undigested.  Kinases  are  also  present  in 
venoms   (Delezeime).     Wchrmann -''  found  that  venom  digests  fibrin 

2"' Arch.  cixp.  Patli.  u.  IMianii.,  100"   (.^fi).  2M);    1011    (('.4),  '244. 
2<i  Univ.  of  Ponii.  Med.  l?ull..  1002    (15),  .'{(JO. 
27  Ann.  d.  I'Inat.  rasteur,  1808   (12),  .510. 


SNAKE  VENOMS  151 

and  inverts  saccharose,  but  does  not  digest  starch.  ^Martin  -^  found 
f.brin  ferments  in  various  venoms,  which  are  probably  important 
aji'ents  in  causing  tlirombosis.  There  are  also  active  lipases  in  ven- 
oms, to  which  many  of  the  effects,  especially  hemolysis  and  fatty  de- 
genei-ation  of  the  tissues,  may  be  at  least  partly  due  (Noguchi),  and 
the  hemolysin  of  cobra  venom  seems  to  be  a  lipase  that  splits  lecithin 
into  hemolytic  substances  (Coca)."^'' 

Toxicity. — Calmette  has  determined  the  toxicity  of  several  venoms, 
and  gives  the  following  tlgures : 

1   crni.  cohra   or  a.s/)i.s  kills 4000  k<iin.  of  rabbit. 

1  grm.  hopluccphaluft  kills 3450      "  "        '' 

1   ^ni.  fer  de  lanre  or  psciidirhis  kills   .       .      SOO       "  "        " 

1  gm.  Crotalus   horridus  kills    ....      600      "  "       " 

1  gm.  Pelias  beriis  kills  .      .      .      .    -  .      .  250      "  "       " 

The  danger  of  the  bite  depends  not  only  upon  the  difference  in 
the  strength  of  the  venom  of  different  varieties  of  serpents,  but  also 
upon  the  size  of  the  snake,  the  time  of  year  and  condition  of  hunger  or 
plenty,  and  particularly  whether  the  entire  discharge  is  injected  suc- 
cessfully or  not.  The  fatal  dose  of  cobra  venom  for  an  adult  man  is 
variously  estimated  at  from  0.01  to  0.03  gm.,  while  the  venom  of  Hy- 
drophiinae  is  about  ten  times  as  toxic ;  for  crotalus  venom  the  lethal 
dose  is  probably  0.15  to  0.3  gm.  (Nogiichi).  Probably  in  the  major- 
ity of  strikes,  by  no  means  all  the  fluid  ejected  by  both  fangs  is  in- 
jected beneath  the  skin  of  the  victim.  A  large  diamond  rattler  may 
eject  as  much  as  a  half  teaspoonful  of  venom  at  one  discharge  and 
such  a  dose  would  usually  be  fatal.  Repeated  ejections  decrease  the 
strength  of  the  venom  rapidly,  until  it  may  have  almost  no  toxicity. 
In  general,  venom  is  most  active  in  w^arm  weather  and  immediately 
after  the  snake  has  fed ;  in  winter  its  toxicity  is  slight. 

The  mortality  in  America  from  snake-bites  is  very  hard  to  ascer- 
tain, various  authors  giving  figures  at  wide  variance.  The  extensive 
studies  of  Willson  -"  show  about  ten  per  cent,  niortalit}-  from  all  venom- 
ous snake-bites  in  this  country^  the  different  species  giving  figures  as 
follows :  Coral  snakes,  twenty  to  fifty  per  cent. ;  water  moccasins, 
seventeen  per  cent. ;  large  rattlesnakes,  eleven  to  twelve  per  cent. ; 
copperheads  and  ground  rattlers,  no  mortality  except  in  children  or  in 
cases  of  complications.  The  mortality  in  children  is  at  least  double 
that  in  adults.  Many  deaths  from  snake-bites  of  all  kinds  are  due 
to  the  treatment  rather  than  to  the  bite.  The  poisonous  snakes  of 
Australia,  although  numerous,  are  not  very  virulent,  and  the  mortal- 
ity is  given  as  about  seven  per  cent.  A  full  charge  of  venom  from 
the  cobra  and  many  other  Indian  snakes  is  inevitably  fatal  (Fayrer). 
The  crotaline  snakes  of  the  tropics  are  more  venomous  than  those  of 

28  .Jour,  of  Phvsiol.,   1905    (32),  207. 
2Sa,Joiir.  Infect.  Dis.,  1015    (17),  351. 

29  Arch.  Int.  Med.,  1008    (1),  516 


152  PHYTOTOXINS    AND    ZOOTOXINS 

the  north,  Lacheris  lanceolatus  of  Central  America  and  Mexico  being 
nearly  as  dangerous  as  the  cobra. 

When  venom  is  taken  into  the  stomach  in  the  intervals  of  diges- 
tion, enough  may  be  absorbed  to  produce  death,  especially  in  the  case 
of  those  venoms  which  contain  a  large  proportion  of  the  albumose, 
which  is  dialyzable ;  but  during  active  digestion  the  venom  undergoes 
alteration  and  is  rendered  harmless.  It  has  been  found  experimen- 
tally in  animals  that  cobra  venom  placed  in  the  stomach  causes  ordi- 
narily no  harm  whatever,  but  if  a  loop  of  the  intestine  is  isolated,  a 
fistula  established  and  allowed  to  heal,  venom  introduced  through 
this  opening  always  produces  death.  It  is  probably  not  so  much 
the  pepsin  and  hydrochloric  acid  that  destroys  the  venom,  as  the 
trypsin.  If  the  bile-duct  is  ligated,  the  venom  is  destroyed  just 
the  same.  Much  of  the  venom  seems  to  be  eliminated  into  the  stom- 
ach, no  matter  how  it  is  introduced  into  the  system,  and  apparently 
it  is  also  partly  excreted  by  the  kidneys.  Rattlesnake  venom  seems 
not  to  be  absorbed  through  mucous  membranes. 

Physiological  Action. — As  indicated  in  the  preceding  paragraph, 
the  effects  of  the  bites  of  different  classes  of  snakes  are  quite  differ- 
ent.    Langmann  describes  the  symptoms  as  follows: 

Cobra  Poisoning. — ^"\'\'itliin  an  hour,  on  an  averajje,  the  first  constitutional 
symptoms  appear:  a  pronounced  vertigo,  quickly  followed  by  weakness  of  the 
legs,  which  is  increased  to  paraplegia,  ptosis,  falling  of  the  jaw  with  paralysis 
of  the  tongue  and  epiglottis;  at  the  same  time  there  exists  an  inability  to  speak 
and  swallow,  with  fully  preserved  sensorium.  The  symptoms  thus  resemble  those 
of  an  acute  bul)>ar  paralysis.  The  pulse  is  of  moderate  strength  until  a  few 
minutes  after  the  cessation  of  respiration ;  the  latter  becomes  slower,  labored, 
and  more  and  more  superficial  until  it  dies  out  almost  imperceptibly.  Death 
occurs  at  the  latest  within  fifteen  hours;  in  32  per  cent,  of  all  cases  in  three 
hours.  There  are  very  few  local  changes."  Cushny  sna  finds  that  cobra  venom 
produces  paralysis  of  the  motor  nerve  terminations  of  muscle,  resembling  the 
action  of  curare ;  the  central  nervous  system  is  not  directly  involved.  Death  re- 
sults from  failure  of  tlie  motor  nerve  ends  in  the  respiratory  muscles  to  transmit 
impulses  to  the  muscles.  Alkaloids  that  are  antagonistic  to  curare  (physostig- 
mine,  guanidine)  are  not  eff"ective  in  cobra  poisoning,  but  are  themselves  rendered 
inactive. 

Viper  Poisoning. — "After  the  bite  of  a  viper  the  local  changes  are  most  pro- 
nounced; there  are  violent  pains  in  the  bleeding  wound,  hemorrhagic  discolora- 
tion of  its  surroundings,  bloody  exudations  on  all  the  mucous  memln-anes,  and 
hemoglobinuria.  Usually  somewliat  later  than  in  cobra  poisoninir  constitutional 
symptoms  develop;  viz.,  great  prostration  with  nansea  and  vomiting,  blood  pres- 
sure falls  continuously,  and  respiration  grows  slow  and  stertorous.  After  a  tem- 
porary increase  in  reflexes,  paresis  supervenes,  with  paraplegia  of  tlie  lower  ex- 
tremities, (>xtending  in  an  upward  direction  and  ending  in  a  comiilete  ))aralysis. 
It  therefore  rescmliles  an  acute  ascending  spinal  paralysis.  If  tlio  patient  re- 
covers from  tlie  paralysis,  a  septic  fccr  may  develop;  not  rarely  there  remain 
suppurating  gangrenous  wounds,  which  heal  poorly." 

It  will  be  noticed  that  there  is  lacking  tlu'  usual  ]ioriod  of  incuba- 
tion that  follows  injection  of  bacterial  toxins,  and  if  it  happens  that 
the  venom  has  been  injected  directly  into  one  of  the  veins,  death  may 

20a  Trans.  Koy.  Soc,  London    (B),   1016    (208),   1. 


NATURE  OF  VENOMS  153 

occur  within  a  few  minutes.  AVhen  recovery  occurs,  the  disappear- 
ance of  symptoms  is  remarkably  abrupt,  within  a  few  hours  a  des- 
perately sick  person  becoming  almost  entirely  free  from  all  evidences 
of  the  intoxication. 

Pathological  Anatomy. — Postmortem  examination  shows  changes  varying  with 
the  nature  of  the  poisonous  shake  that  has  caused  death.  In  the  case  of  a  cohra 
bite,  according  to  JMartin.  the  areolar  tissue  aVmut  tlie  wound  is  infiltrated  with 
pinkish  fluid;  the  blood  is  often  lluid;  the  \eins  of  the  jiia  are  congested,  and 
the  ventricles  often  contain  turbid  lluid;  the  kidneys  inay  show  much  congestion. 
When  death  occurs  in  a  few  minutes,  enormous  general  intravascular  clotting  is 
found,  which  seems  to  be  the  cause  of  death.  After  death  from  a  viper  bite  the 
site  of  the  wound  is  the  seat  of  intense  edema  and  extravasation  of  blood;  if  in 
the  muscles,  these  are  much  softened  and  disorganized.  Hemorrliages  are  fovind 
in  all  organs  and  in  the  intestinal  tract.  If  death  occurs  after  several  days  it 
is  generally  because  of  sepsis,  and  shows  the  usual  changes  of  this  condition ;  in 
addition,  as  a  rule,  to  marked  gangrenous,  ulcerative,  and  sloughing  processes  at 
the  site  of  the  bite. 

Histologically  there  are  found,  in  addition  to  innumerable  hemorrhages  in  nearly 
all  the  organs,  many  vessels  plugged  with  thrombi  composed  of  more  or  less 
hemoh'zed,  agglutinated  erythrocytes.  The  changes  produced  in  tlie  nervous  tissue 
by  the  Australian  tiger  snake  are  described  by  Kilvington.so  ^vho  foimd  marked 
chromatolysis,  the  Nissl  bodies  breaking  into  dust-like  particles,  and  eventually 
all  stainable  substance  disappearing  from  the  cytoplasm;  the  nucleus  retains  its 
central  position,  but  often  loses  its  outline  and  may  disappear.  The  cells  aroimd 
the  central  canal  of  the  cord  are  most  affected.  There  are  no  inflammatory 
changes  in  the  nervous  system,  and  if  death  occurs  very  quickly  there  may  be  no 
microscopic  alterations.  Hunter  si  found  similar  changes  in  the  Nissl  bodies  in 
both  krait  and  cobra  poisoning:  in  the  medullated  fibers  he  found  the  myelin 
sheath  converted  into  ordinary  fat.  The  venom  of  sea  snakes  (Enhijdrina  valaka- 
dien)  has  a  severe  action  on  the  nervous  tissues,  while  Daboia  has  none  (Lamb 
and  Hunter  32).  Nowak  33  studied  experimental  animals,  and  found  mucli  fatty 
change  in  the  livers,  even  if  death  occurred  one-half  hour  after  poisoning;  also 
focal  necrosis  in  the  liver,  acute  parenchymatous  alterations  in  the  kidney,  and 
pneumonic  patches  in  the  lungs. 

Effects  on  the  Blood. — Tlicre  has  been  much  discussion  concerning  the  part 
played  by  tlie  abvnidant  and  prominent  intravascular  clotting  in  causing  death 
after  snake-bite.  Lamb  s*  states  that  when  venoms  are  slowly  absorbed  the 
coagulability  of  the  blood  is  decreased  and  it  is  found  fluid  after  death,  but  when 
a  fatal  dose  of  venom  (viper)  is  rapidly  absorbed,  clotting  is  increased  and  throm- 
bosis is  the  chief  cause  of  death.  ]\Iartin  has  demonstrated  very  active  fibrin 
ferments  in  snake  venom  (loc.  cit.).  It  is  highly  probable,  however,  that  many 
of  the  thrombi  of  venom  poisoning  are  not  produced  by  coagulation  of  fibrin,  but 
by  agglutination  of  the  red  corpuscles,  which  Flexner  35  has  shown  can  cause 
large  clots  in  the  heart  and  great  vessels,  as  well  as  "hyalin"  thrombi  in  the 
small  vessels. 

Nature  of  Venoms. — The  varied  effects  produced  by  venoms  have 
been  found  to'  be  due  to  a  number  of  poisonous  elements  which  they 
contain,  and  which  have  been  distinguished  and  separated  from  one 
another  by  Flexner  and  Nognchi.^''  These  are  hemotoxins  (hemoly- 
sins and  hemagglutinins) ,  leucocytolysins,  neurotoxins,  and  endothel- 

30  Jour,  of  Physiol.,  1902  (28),  420. 

31  Glasgow  Med.  -Jour.,  1003    (59),  98. 

32  Lancet,  1907    (ii),  1017. 

33  Ann.  d.  I'Inst.  Pasteur,  1898  (12),  369. 

34  Indian  Medical  Gazette,  Dec,  1901. 

35  Univ.  of  Penn.  Med.  Bull..  1902   (15).  324 

36  Jour.  Exp.  Med.,  1903    (9),  257;  Univ.  Penn.  Med.  Bull.,  1902    (15),  345. 


154  pnyTOToxi\i<  axd  zootoxixs 

iotoxins  (hemorrhagin) ,  but  it  must  be  taken  into  consideration  that 
Faust  ^'  believes  tliat  the  single  o-lucosidal  poison  which  he  has  found 
in  rattlesnake  venom  is  responsible  for  all  the  effects  of  the  venom, 
except  the  hemao-glutination.  [In  another  place  (see  "Hemolysis") 
the  nature  of  the  hemolytic  agent  is  discussed.]  Venom  agglutinin  is 
quite  independent  of  the  hemolysin,  for  it  is  destroyed  by  heating  to 
75°-80°,  whereas  the  hemolysin  is  destroyed  only  partly  at  100°. 
Agglutinin  acts  in  the  absence  of  serum  complement,  and  therefore 
is  not  an  amboceptor;  it  is  apparently  more  like  the  toxins  in  its  na- 
ture. The  agglutination  of  the  corpuscles  does  not  interfere  with 
their  subsequent  hemolysis.  Michel  states  that  the  agglutinin  of  cobra 
venom  can  be  separated  from  the  hemolysin  and  the  toxin  by  means  of 
ultrafiltration  through  collodion  membranes,  as  the  agglutinin  exists 
in  larger  molecular  aggregates.^*'-'' 

The  leucocytotoxins  were  found  by  Flexner  and  Noguchi  to  be 
quite  distinct  from  the  hemolysins,  for  after  saturating  all  the  hemoly- 
sin with  red  corpuscles,  the  venom  still  shows  its  efit'eets  on  the  leu- 
cocytes, which  effects  consist  in  cessation  of  motility  and  disintegra- 
tion, affecting  particularly  the  granular  cells.  The  leucocytotoxin, 
however,  resembles  the  homolysin  in  that  it  appears  to  be  an  ambo- 
ceptor. Leucocytes  are  also  agglutinated  by  venom,  possibly  by  the 
same  agglutinin  that  acts  on  the  red  corpuscles.  Serum  complement 
is  inactivated  in  vitro  by  cobra  venom  through  changes  in  the  globu- 
lins brought  about  by  the  venoms.^^'' 

By  saturating  venom  with  either  red  corpuscles  or  nerve-cells  it 
was  found  by  these  authors  that  the  toxic  principle  for  each  is  dis- 
tinct and  separate.^*^  Other  sorts  of  cells,  however,  are  able  to  com- 
bine, or  at  least  remove  some  parts  of  the  toxic  elements,  but  to  a 
much  less  degree.  The  neurotoxin,  like  the  hemolysin,  resembles  an 
amboceptor,  and  since  venom  contains  no  complement,  the  neurotoxin 
has  first  to  be  supplied  with  complement  by  the  victim's  blood  or  tis- 
sues before  it  can  harm  the  cells.  The  venoms  are  not  only  toxic  for 
mammalian  cells,  but  also  for  simple  unicellular  organisms,  including 
bacteria;  tadpoles  are  paralyzed  in  solutions  containing  one  part  of 
cobra  venom  per  million.^" 

The  pronounced  hemorrhage-producing  projjcrty  of  serums,  partic- 
ularly that  of  the  rattlesnake,  was  also  found  to  be  due  to  a  specific 
toxin  acting  on  the  endothelium  of  the  capillaries  and  small  veins,  and 
not  to  the  changes  in  the  blood  itself,  as  had  formerly  been  thought. 
This  endotheliotoxin,  which  Plexiu^r  and  Noguchi  call  "hemorrhagin," 
is  quite  distinct  from  the  other  toxic  substances,  being  destroyed  at 

37  Arch.  Expor.  Patli.  ii.  I'lunni.,  1!)11    (f>4),  244. 
3«a  Conipt.  Itoiul.  Soc.  ]?ioI.,  l!)lf)   (77).  ISO. 
37anirsclif<'Id  and  Klinj^or.  Ilioohem.  Zcit.,   101;i    (70).  .SOS. 

38  r5an<r  aiuipvcrton  slate  tliat  corpuscles  can  take  up  tlie  uoiroloxiu.  wliicli 
is  soluble  ill  ^<<Ts  and  li])oids. 

30  Ban;:  aiul-^veiton,   I'.ioclieni.   Zeit..    I'Hl     (.SI),   24:?. 


VARIATIONS  IN  VENOMS  155 

75^,  H  tciniK'i'atiirc  that  leaves  tlie  neurotoxin  and  lieiiiolysin  uiiiii- 
jured.  Its  endotlieliolytic  action  is  shown  in  the  glomerular  capil- 
laries, where  it  causes  hemorrhage  and  hematuria  (Pearce)."' 

Variations  in  Venoms. — In  distribution  among  the  various  poison- 
ous reptiles  these  toxins  seem  also  quite  distinct  from  one  another, 
which  explains  the  difference  in  the  et't'ects  of  bites  b}'  snakes  of  various 
kinds.  Cobra  venom  contains  chiefly  neurotoxin,  hence  the  symp- 
toms of  cobra  bite  are  largely  of  nervous  origin,  with  but  little  local 
tissue  change.  Rattlesnake  venom  owes  its  effects  chiefly  to  hemor- 
rhagin,  hence  the  marked  local  necrosis  and  extravasations  of  the 
blood,  and.  the  generalized  hemorrhages ;  the  nervous  effects  following 
viper  bite  are  probably,  in  part,  due  to  hemorrhages  in  the  nervous 
tissue.  Cobra  venom  produces  great  hemolysis  and  little  agglutina- 
tion. Rattlesnake  venom  has  relatively  little  agglutinative  or  hemo- 
lytic power.  Water  moccasin  and  copperhead  venoms  are  more  ag- 
glutinative than  either,  and  intermediate  in  hemolytic  strength;  they 
cause  much  local  tissue  destruction. 

The  exact  action  of  cobra  venom  on  various  centers  and  orfj^ans  has  been 
studied  by  Elliot. 4i  It  raises  blood  pressure  when  in  dilution  of  1  :  10,000,000, 
by  contractinjj  vessels  and  stimulating  the  Iieart;  low  lethal  doses  kill  by  para- 
lyzing the  respiratory  center. 

Krait  {Bunf/arus  c(rrulites)  venom  acts  similarly,  but  less  powerfully,  and 
cannot  be  neutralized  by  Calmette's  antivenin.^2 

Sea-snake  venoms  are  by  far  the  most  poisonous  of  all.  For  Enhj/drina  valalca- 
dien  the  lethal  dose  for  rabbits  is  0.00006  gram  per  kilo  body  weight.  It  acts  by 
vaaus  stimulation  and  paralysis  of  respiratory  centers  and  of  motor  nerve- 
endings.43 

Russell's  viper  (Dahoia  T'lissellii)  owes  its  effects  chiefly  to  intravascular  clot- 
ting, according  to  Lamb  and  Hanna,**  and  contains  no  neurotoxin.  It  is  not 
neutralized  l)y  Calmette's  antivenin.  The  clots  are  due  to  agglutination  and  con- 
tain no  fibrin    (Flexner). 

The  "Gila  Monster"  {Heloderma-  suspectum)  seldom  causes  serious  poisoning 
in  man,  but  may  kill  small  animals,  such  as  frogs. -is  Its  poison  is  only  slightly 
hemolytic,  but  produces  degenerative  changes  in  the  nervous  system  (Langniann). 
The  hemolysin  is  activated  by  lecithin  (Cooke  and  Loeb).  An  elaborate  series  of 
studies  by  Leo  Loeb  and  his  associates  give  all  the  known  facts  concerning  the 
Gila  Monster.45a 

Loss  of  Bactericidal  Powers. — The  frequency  of  marked  and  per- 
sistent sloughing  and  suppuration  at  the  site  of  snake-bites,  particu- 
larly from  the  vipers,  and  the  common  termination  in  sepsis,  was 
attributed  by  Welch  and  Ewing  *^  to  a  loss  of  bactericidal  power  of 

4o,T(nir.  Exper.  Med.,  1009   (II),  .5.32. 

41  Lancet,  1904   (i),  715. 

42  Elliot,  Sillar,  and  Carmichael,  Lancet,  1904    (ii),  142. 

43  Eraser  and  Elliot,  Lancet,  1904  (ii),  141;  also  Rogers,  Jour,  of  Physiol., 
190.3  (30),  iv.  The  above  are  also  given  completely  in  the  Philosophical  Trans- 
actions of  the  Roval  Societv,  1904-5,  vol.   187. 

44, Jour,  of  Patli.  and  Bact.,  1902   (8),  1. 

45  Thorough  studv  bv  Van  Denburgli  and  Wright,  Amer.  .Toin\  of  Plivsiol., 
1900    (4),  209. 

45a  Carnegie  Inst.  Publication  Xo.  177,  191.3. 

46  Lancet,  1894  (1),  1236;  Ewing,  ]\Ied.  Record,  1894   (45),  66,3. 


156  PHYT0T0XIK8    AND    ZOOTOXINS 

the  blood,  which  they  found  followed  experimental  venom  poisoning. 
This  has  been  ascribed  by  Flexner  and  Noguchi  to  saturation  of  serum 
complement  by  the  numerous  amboceptors  of  the  venoms,  so  that  no 
complement  is  left  for  the  serum  to  use  against  the  bacteria.  In 
senim  whose  complements  do  not  combine  with  the  venom  amboceptors 
(e.  g.,  Necturus)  the  normal  bactericidal  powers  are  not  in  the  least 
impaired  by  the  addition  of  venom.  Morgenroth  and  Kaya  ascribe 
the  loss  of  complement  to  a  destruction  by  some  agent  in  the  venom. 

Snake  Serum. — The  serum  of  serpents  is  also  toxic  for  other  ani- 
mals,*®'' even  when  tlie  serpent  is  not  a  venomous  one ;  e.  g.,  the  harm- 
less pine  snake  (Pityophis  catenifcris).  The  toxicity  of  snake  serum 
seems  to  depend  chiefly  upon  its  hemotoxic  effects  (hemagglutination 
and  hemolysis),  the  toxic  substances  resembling  amboceptors  and  sim- 
ilar to,  but  not  altogether  id'entical  with,  the  amboceptor  of  the  ven- 
oms. Crotalus  tissues  also  produce  poisoning  in  proportion  to  the 
blood  they  contain,  but  are  without  toxic  effects  of  their  own  (Flex- 
ner and  Noguchi). 

Antivenin. — Snake  venom  has  the  essential  propertj^  of  all  true 
toxins  of  immunizing,  with  the  appearance  of  an  antitoxin  in  the 
blood.  The  first  successful  immunizations  seem  to  have  been  made 
by  Sewall,*"  but  the  practical  production  of  antitoxic  serum  was  first 
accomplished  by  Calmette  *^  and  by  Fraser.*^  At  first  it  was  be- 
lieved that  cobra  antivenin  neutralizes  the  neurotoxins  and  hemoly- 
sins of  venoms  of  any  origin,  and  also  of  snake  serums,  and,  therefore, 
should  be  quite  effective  against  cobra  and  similar  venoms  which  pro- 
duce chiefly  neurotoxic  and  hemolytic  changes.  This  implies  that  these 
toxic  substances  are  of  identical  nature  in  all  snakes,  no  matter  how 
dissimilar  the  snakes  may  be,  but  various  investigators,  especially 
Lamb,  have  found  sufficient  specificity  exhibited  by  different  venoms 
and  antivenoms  to  indicate  the  necessity  of  employing  the  specific 
antiserum  in  each  case  of  snake  bite.  A  special  antitoxin  against  rat- 
tlesnake venom  and  its  hemorrhagic  toxin  has  been  successfully  pre- 
pared by  Noguchi.-'^*'  This  crotalus  antivenin  also  neutralizes  hemo- 
lysins of  various  venoms,  and  also  of  snake  serums. 

Presumably  antivenin  neutralizes  venoms  in  exactl.y  the  same  way 
that  antitoxin  neutralizes  toxins;  i.  e.,  cell  receptors  are  tlirown  off 
from  the  injured  cells  during  immunization,  which  combine  with 
venom  amboceptors  in  the  blood,  and  thus  prevent  their  combining 
vnth  the  cells.  Antivenin  also  prevents  the  inhibiting  action  of  venom 
on  bactericidal  serum,  indicating  that  it  prevents  the  venom  ambo- 
ceptors from  binding  the  serum  comj^h'iuciit.     Tlie  reaction  of  venom 

40a  Questioned  hv  Wolkcr  and  :\rarsliall,  Jour,  riiannacol..  1015    (0),  503. 
47  Jour,  of  Pliysiol.,  ]SS7   (S).  203. 

48i\nn.  d.  I'Insl.  rastcur,  18!)4  (6),  275;  also  sul)so(iiu'n(  arfu-les  in  1897  (11), 
214;    1808    (12),   .343. 

49  British  Mod.  Jour.,  1805    (i).  1300. 

coUniv  of  I'.Miii.  Med.  I'.iill..  1004    (17),  154;  Joiir.  Exper.  Mod.,  1006    (8),  G14. 


SCORPION  POISON  157 

and  antiveiiin  is  certainly  a  chemical  one,  being  likened  by  Kyes  ^^ 
to  that  of  strong  acids  upon  strong  bases. 

The  serum  of  animals  immunized  to  venoms  contains  precipitins 
for  the  proteins  of  these  venoms,  and,  to  some  extent,  for  the  serum 
proteins  of  the  same  species  of  snakes.  These  precipitins  are  highly 
but  not  absolutely  specific,  and  thc}^  bear  no  exact  quantitative  rela- 
tion to  the  other  antibodies  present  in  the  same  sera. 

As  is  well  known,  snakes  are  nearly  or  quite  insusceptible  to  snake 
venom.  Cunningham  ^-  found  that  serum  of  cobras  was  devoid  of 
antitoxic  property,  so  the  immunit3'  of  snakes  must  be  ascribed  to  an 
absence  of  cell  receptors  in  their  tissues,  with  which  their  venom  am- 
boceptor can  combine.  The  reputed  immunity  of  the  mongoose  and 
hedgehog  depends  partly  on  a  relatively  low  susceptibility,  but  prob- 
ably more  on  the  agility  of  the  mongoose  and  the  defensive  spines  of 
the  hedgehog. 

Platypus  Venom. — The  only  mammal  with  a  venomous  secretion 
is  that  strange  freak,  the  duck-billed  platypus  (Ornithorhynchus  par- 
adoxus). The  males  have  a  hollow  movable  spur  on  each  hind  foot, 
communicating  like  a  fang  with  the  venom  gland,  which  secretes  a 
venom  with  properties  resembling  the  venoms  of  the  Australian  snakes, 
but  much  weaker. 

SCORPION  POISON -3 

This  poison  is  secreted  by  a  pair  of  specialized  glands  in  the  pos- 
terior segment  of  the  elongated  abdomen,  surrounded  by  a  firm  cap- 
sule with  a  sharp  apex  through  which  the  poison  is  discharged.  Its 
efll'ect  on  man  is  usually  confined  to  local  pain,  swelling,  and  occa- 
sionally phlegmonous  inflammation  with  constitutional  s^'mptoms 
after  bites  from  the  largest  species.  In  Africa  a  large  scorpion  (An- 
droctonus)  exists,  that  is  reputed  frequently  to  cause  fatal  poisoning, 
especially  in  children.  The  majority  of  serious  results  folio-wing 
scorpion  bites,  as  well  as  bites  of  poisonous  insects  to  be  considered 
later,  are,  however,  due  to  infection  of  the  wound,  which  occurs  read- 
ily because  of  local  necrosis  and  hemorrhages,  and  also  because  of 
the  unfavorable  conditions  existing  in  tropical  climates.  Apparently 
these  bites  favor  local  infection  much  as  do  those  of  vipers. 

When  general  symptoms  do  occur,  they  are  described  as  resembling 
strychnine  poisoning,  with  trismus,  stiffness  of  the  neck  and  eventu- 
ally of  the  respiratory  muscles,  which  seems  to  be  the  chief  cause  of 
death  (Cavorez).  Thompson,'^*  however,  observed  only  seldom  severe 
symptoms,  consisting  of  general  paralysis  that  passed  off  in  a  few 
hours.     Most  experimenters  with  scorpion  poison  describe  it  as  chiefly 

r.iBerl.  klin.  Woch.,  1904    (41),  494. 

52Xature,  1896    (5.5),  139. 

53  A  complete  discussion  of  the  literature  on  poisonous  invertebrates,  etc.,  is 
given  by  v.  Fiirth,  "Vergleichende  cliemiselie  Phvsiologie  der  niederen  Tiere," 
Jena.  1903:  and  bv  Faust,  "Die  tierischen  Gifte,"  Braunsclnveig.  190G. 

54Proc.  Acad.  Nat.  Sci.  of  Philadelphia,  1S8C,  p.  299. 


158  PnYTOTOXIXS   axd   zootoxixs 

a  nerve-tissue  poison,  and  it  also  seems  to  act  as  a  hemolysin  and  ag- 
glutinin (Bellesme  and  Sanarelli),  but  Todd'"'  found  it  without  ac- 
tion on  corpuscles  and  not  capable  of  combining  with  nervous  tissues. 
Calmette  '■•"  gives  the  lethal  dose  for  a  guinea-pig  as  0.5  milligram, 
while  Phisalix  and  Varigiiy  put  it  at  0.1  milligram  and  state  that 
scorpion  blood  is  also  poisonous.  Wilson  "  found  the  toxicity  of  the 
venom  equal  to  0.1  gram  per  million,  that  is,  one  gram  of  poison  will 
kill  10,000,000  grams  of  guinea-pig,  hence  it  is  much  stronger  than 
cobra  venom.  It  is  quite  stable,  and  keeps  many  months  in  an  ice 
chest;  is  not  affected  by  heating  to  100°  for  a  brief  period,  but  is  de- 
stroyed after  12  or  13  minutes'  heating.  The  active  constituents  are 
precipitated  by  saturating  with  ammonium  sulphate,  or  by  an  excess 
of  alcohol.  The  average  amount  of  toxin  in  an  Egyptian  scorpion 
{Buthus  quinque  striatus)  is  sufficient  to  kill  about  35  kilos,  which 
agrees  with  the  fact  that  fatal  poisoning  by  this  scorpion  is  rare  in 
adults,  but  reaches  60  per  cent,  in  children.  The  venom  is  harm- 
less when  taken  into  the  stomach,  and  is  said  to  be  made  inactive  by 
ammonia,  calcium  hypochlorite,  and  peroxide  of  hydrogen.  Calmette 
claims  that  antivenin  for  cobra  in  part  neutralizes  scorpion  poison,  a 
statement  which  could  not  be  corroborated  by  Todd,  who  succeeded, 
however,  in  preparing  an  efficient  antiserum  by  immunizing  horses 
with  scorpion  venom.  A  large  number  of  naturalists  and  raconteurs 
have  furnished  interesting  tales  of  suicide  by  scorpions,  which  are 
more  than  improbable  in  the  light  of  our  present  knowledge  concerning 
natural  immunity.  Many  animals  seem  to  possess  more  or  less  im- 
munity to  scorpions  (AVilson),  especially  such  wild  animals  as  are 
much  exposed  to  them. 

SPIDER  POISON 

The  poison  apparatus  of  the  spiders  consists  of  two  long  pouches 
lying  in  the  thorax  and  extending  into  the  jaws,  at  the  apex  of  which 
the  poison  is  discharged.  Some  of  the  larger  members  of  the  family 
are  very  poisonous,  e.  g.,  the  Malmignatte  {Lathrodectes  tredecim- 
guttatas),  of  the  vicinity  of  the  lower  Volga  in  southern  Russia,  is 
said  to  have  destroyed  70,000  cattle  in  one  year,  the  bite  being  fatal 
in  12  per  cent,  of  all  cases,  although  rarely  killing  man.  Other 
members  of  this  species  in  Chili,  ^Madagascar,  and  other  countries 
are  not  much  less  venomous.  Kobert  has  studied  the  poison  of  Malmi- 
gnatte and  found  it  distributed  throughout  the  body  of  the  spider, 
even  in  the  eggs,  and  resembling  in  nature  the  snake  venoms.  It  is 
destroyed  by  heating,  and  seems  to  be  of  protein  nature;  the  chief 
effect  is  u])()n  llie  nervous  system  and  heart. '^ 

f.r.  Jour,  of  Tlvtriono,   1000    (9),  fif). 
r.n  Ann.  Inst.  "Pastour,    ISO.'i    (9),   2.32. 

5 T  Records  of  Kfr\i)tiini  (iov'1..  School  of  Med..  1!)04:  iibsl.  in  Jonr.  of  riiysiol., 
1904    i'M),  p.  xlvii'i 

•"'R  In    wcstcin    Aiiicricii    is    fo\iii(l    ;i    snidiT     ( l,(i  t  rodcrtcs    iiuictnns)     lli(>    liite    of 


CENTIPEDES  159 

A  number  of  common  spiders  investigated  by  Kobert  ^^  were  ap- 
parently not  poisonous  for  mammals,  except  the  ''cross  spider"  {Epe- 
ira  diadema),  which  has  since  been  thoroughly  studied  by  him  and  by 
Sachs.'^*'  Walbum  *'°*  states  that  the  chief  poison  of  these  spiders  is 
found  in  the  ovaries,  the  salivary  poison  being  much  weaker,  and  the 
hemolysin  is  found  chiefly  in  the  albumin  fraction.  Epeiratoxin  re- 
sembles the  snake  venoms  strikingly,  according  to  Sachs,  for  it  eon- 
tains  a  powerful  hemolysin  which  he  calls  "  arachnolysin, "  acting  very 
differently  with  different  sorts  of  blood,  and  destroj'ed  by  heating  at 
70^-72°  for  forty  minutes,  and  it  behaves  with  lecithin  and  cholesterol 
like  cobra  venom. "^  The  agglutinin  is  quite  distinct  from  the  hemo- 
lysin."- Only  such  blood  is  hemolyzed  as  is  able  to  bind  the  poison 
in  the  stroma  of  the  red  corpuscles.  By  immunizing  a  guinea-pig 
Sachs  succeeded  in  securing  an  antitoxin  of  some  strength.  The  dis- 
covery of  this  hemolysin  explains  Robert's  observation  of  hemoglobin, 
methemoglobin,  etc.,  in  the  urine  of  persons  bitten  by  spiders. 

Von  Fiirtli  considers  that  the  bite  of  the  historically  famous  Italian 
tarantula  is  able  to  cause  no  more  than  local  inflammation,  and  Ko- 
bert found  that  the  entire  extract  of  six  Russian  tarantulas  (w^liich 
are  supposed  to  be  more  poisonous  than  the  Italian)  caused  no  symp- 
toms when  injected  into  a  cat. 

In  all  probability  the  other  poisonous  spiders  possess  toxic  sub- 
stances allied  to  those  of  the  venoms,  with  hemolytic,  agglutinative, 
and  neurotoxic  products,  Sachs'  studies  indicating  the  general  sim- 
ilarity of  all  the  zootoxins.  An  antitoxin  is  said  to  have  been  secured 
against  the  Russian  tarantula."^ 

CENTIPEDES 

Undoubtedly  the  severity  of  centipede  poisoning  has  been  greatly 
exaggerated,  the  results  being  usually  limited  to  local  inflammation, 
frequently  spreading  some  distance  in  an  erysipelas-like  manner. 
An  authentic  case  of  fatal  poisoning  of  a  child  four  years  old  by  a 
centipede  (Scolopendra  heros)  has  been  reported  from  Texas  by  G. 
Linceicum,*'^  death  resulting  five  to  six  hours  after  the  bite  was  re- 
ceived. Besides  the  local  pain  and  inflammation,  vomiting  was  marked, 
occurring  also  in  five  other  non-fatal  cases. 

Centipedes  secrete  their  poison   in  relatively  large   glands,   which 

which  is  capable  of  causing  severe  spasm  of  Uie  abdominal  muscles,  according  to 
Atwood  (Southern  Californ.  Pract..  Vols.  10.  12  and  16).  Kellogg  and  Coleman 
(.Jour,  of  Parasitol.,  1015  (1).  IflJ),  found  extracts  of  the  poison  glands  of  this 
spider  to  be  highly  toxic. 

59  "Beitriige  ziir  Kentnisse  der  Ciftspinnen,"'  Stuttgart.  1001. 

eoHofmeister's  Beitr..   1902    (2),  12.5. 

GoaZeit.  Tmmunitiit.,  1015    (2.3),  623. 

«iPini.  II  Policlinico    (Sez.  Med.),  1009    (16),  208. 

62  V.  Szily,  Zeit.  Tmmunitiit.,  1910   (.5).  2S0. 

<5»  Konstanzoff,  Russkv  Wratsch.,  1007,  Xo.  17. 

64Amer.  Jour.  Med.  Sci.,  1866   (.52),  575. 


160  PHYTOTOXIXS    AND    ZOOTOXINS 

discliarge  at  the  apices  of  a  pair  of  specialized  claws  that  take  the 
place  of  the  first  pair  of  legs.  The  nature  of  this  poison  seems  not 
to  have  been  investigated.  Numerous  chemical  substances  are  de- 
scribed as  secreted  by  other  glands  of  these  animals,  including  prus- 
sic  acid  and  a  camphor-like  matter  (see  v.  Fiirth). 

BEE  POISON 

Bee  poison  has  been  better  studied  than  most  insect  poisons,  begin- 
ning with  the  work  of  Paul  Bert  (1865).  It  is  secreted  by  the  glands 
into  a  small  poison  sac,  and  stored  up  until  ejected.  Cloez  found 
that  bee  poison  was  precipitated  by  ammonia,  tannin,  and  platinic 
chloride,  and  Langer  proved  it  to  be  a  non-volatile  organic  base.  As 
excreted,  it  is  acid,  contains  30  per  cent,  of  solids,  and  one  honey-bee 
secretes  0.0003-0.0004  gm.  It  contains  formic  acid  and  much  pro- 
tein, but  it  has  been  stated  that  the  poison  is  protein-free,  and  is  not 
destroyed  by  heat  (100°),  weak  acids,  or  alkalies.  On  the  other  hand, 
it  is  said  to  be  destroyed  by  proteolytic  enzymes,  which  would  indicate 
that  it  is  of  protein  nature.  Hemolysis  is  produced  both  in  v-itro 
and  in  vivo  with  all  sorts  of  blood,  but  to  very  different  degrees, 
thus  resembling  spider  toxin.  The  hemolytic  action  is  greatly  in- 
creased by  the  presence  of  lecithin,  forming  a  toxolecithid  like  "cobra 
lecithid. " ''^  Locally  bee  poison  causes  necrosis,  with  marked  hyper- 
emia and  edema.  A  4500  gm.  dog  was  killed  by  intravenous  injection 
of  6  c.c.  of  a  1.5  per  cent,  solution  of  pure  poison  (Langer).*'*' 

Immunity  is  undoubtedly  possible,  for  bee-keepers  frequently  show 
a  great  decrease  in  susceptibility.  On  the  other  hand,  abnormally 
great  susceptibility  is  frequently  seen,  some  cases  of  fatal  poisoning 
having  been  obsei'ved.*''^ 

Wasps  and  Hornets  presumably  produce  poisons  similar  to  those  of 
the  bees.  A  study  by  Bertarelli  and  Tedeschi  '^'"^  establishes  this  for  a 
species  of  wasp  (Vespa  crahro  L) . 

Ants  also  produce  formic  acid,  a  fact  so  well  known  that  it  has 
come  to  be  considered  that  this  is  the  source  of  their  toxicit.y.  Von 
Fiirth,  however,  suggests  the  probability  that  ant  poison,  like  that  of 
the  bees,  owes  its  chief  effects  to  other  more  complex,  unkno\vn  poi- 
sons.*'^ 

«5  Mor<jenroth  and  Carpi.  Borl.  klin.  Wocli.,  lOOfi    (43).   1424. 

66  Arch.  exp.  Path.  u.  Pharm.,  1890  (,3S).  .3.S1;  Arcli.  intornat.  Pliannac.  ot 
Ther.,  1800   (6),  181. 

«7  Hospitalstidcndp,  100,5,  No.  27. 

«7aCVnt.  f.  Pakt..   101,3    (68).  .300. 

fisAn  at.tompt  by  P.arratt  (Ann.  Trop.  IVfcd.  and  Parasit(,l..  1010  (4).  177)  to 
olitain  a  poison  from  culex  mosqnitos  was  nnsnooossful.  Tlio  Iwdios  of  "black 
flios"  contain  an  activo  poison  tliat  could  not  bo  idontifiod  by  Rtokos  (,Tonr.  C\it. 
Dis.,  1014  (32),  8,30),  beyond  thai  it  is  insolul)lo  in  alcohol,  which  docs  not 
inactivate  it,  and  that  it  is  destroyed  by  trypsin. 


TOADS  .\.\J>  SALAMAXDERS  161 


POISONS  OF  DERMAL  GLANDS  OF  TOADS  AND  SALAMANDERS 

It  has  been  known  for  centuries  that  toads  produce  poisonous  sub- 
stances. Pare  in  1575  havin<r  discoursed  interestingly,  if  inaccurately, 
on  this  topic.  Numerous  studies  have  been  made  of  these  poisons, 
which  are  secreted  by  the  dermal  glands  and  therefore  cannot  be  used 
for  poisoning  either  prey  or  enemies  (except  those  that  feed  upon 
them)  ;  the  most  extensive  study  being  that  of  Faust.*'''  He  isolated 
two  constituents,  ai)i)arently  the  same,  in  different  species  of  toads; 
one,  which  he  called  hufotalin,  is  very  active,  resembling  the  digitalis 
group;  the  other,  bufonin,  is  much  less  active.  Bufonin  is  neutral  in 
reaction,  soluble  in  warm  alcohol,  but  slightly  in  cold.  Analysis  in- 
dicates an  empirical  formula  of  C34H-4O0.  It  probably  is  the  cause 
of  the  milky  appearance  of  the  dermal  secretion.  P)ufotalin  seems  to 
be  C34ll4eOio,  is  acid  in  reaction,  soluble  in  chlorofonn  and  alcohol, 
but  not  in  petroleum  ether.  Subcutaneous  injection  of  2.6  mg.  bufo- 
talin  killed  a  dog  (weighing  4  kg.)  in  four  to  five  hours;  given  by 
mouth  it  causes  much  vomiting  and  diarrhea,  so  that  large  doses  are 
not  fatal.  It  causes  much  local  irritation  when  applied  to  mucous 
membranes,  but  produces  no  marked  changes  at  the  site  of  injection. 
The  effects  on  the  circulation  resemble  in  all  respects  those  of  the  digi- 
talis group ;  bufonin  acting  similarly  but  much  weaker  than  buf otalin. 
Fiihner  ^°  considers  bufotalin  to  be  more  closely  related  to  the 
saponins.  Bufotalin  seeins  to  be  derived  from  bufonin  by  oxidation, 
and  the  latter  is  quite  similar  to  cholesterol,  apparently  having  the  fol- 
lowing formula:  IlO-}I.,QC-^.j~C^^lIn,-OH.  An  important  consider- 
ation is  that  Faust  has  also  isolated  from  the  venom  of  cobra  and  cro- 
talus,  poisons  which  seem  related  to  these  toad  poisons,  the  cobra  poi- 
son being  assigned  an  empirical  formula  of  C-^^^H^g^^^q,  and  the  cro- 
talus  poison  C.^^H-^Ooi. 

Phisalix  and  Bertrand  "^  have  found  poison  in  the  blood  of  toads 
similar  to  that  of  the  glands.  The  hemolytic  property  observed  by 
Pugliese  '^^  may  be  due  to  the  acidity  of  the  dermal  secretion.  The 
poisons  of  different  species  seem  to  be  quite  the  same  in  all  (Faust). 
From  the  dermal  secretion  of  the  large  tropical  toad,  Bufo  agua, 
Abel  and  Macht  ^^  have  isolated  two  distinct  active  substances ;  one 
identical  with  epinephrine,  which  constitutes  nearly  seven  per  cent,  of 
the  crude  venom;  the  other,  which  makes  up  36  per  cent.,  is  called 
hufagin,  has  a  composition  indicated  by  the  formula  C-i^Ho^Os,  and 
therefore  is  presumably  related  to  the  rest  of  this  group  which  arises 
from  cholesterol.     In  physiological  action  bufagin  resembles  digitalis, 

69  Arch.  f.  exp.  Patli.  u.  Pharra.,   1902    (47),  279.     Complete  bibliography  and 
review. 

70  Arch.  exp.  Path.  11.  Pharni..   1910    (6.3),  374. 

71  Arch.  d.  physiol.  norm,  ot  path.,   1S93    (5),  511. 

72  Arohivio  di  farm.  e.  terap.,  1894  (2),  321;  Arch,  ital  de  Biol..  1S95    (22),  79. 

73  Jour.  Amer.  Med.  Assoc,  1911   (")(>),  1.531;  Jour,  of  Pharm.,  1912   (3),  319. 

11 


162  PHYTOTOXINS    AXD    ZOOTOXiys 

and  it  is  extremely  active.  The  toad  is  relative!}^  immune  to  bufagiu, 
but  not  at  all  to  the  epinephrine.  A  Chinese  drug  derived  from  toad 
skins  has  been  found  to  contain  similar  ing-redients  (Shimizu"^),  as 
well  as  a  substance  resembling  picrotoxin  in  action. 

Salamanders  also  produce  poisonous  secretions  in  their  dermal 
glands,  which  have  been  studied  especially  b}^  Faust,'*  and  earlier 
by  Zalesk}^,'^^  who  isolated  an  organic  base  which  he  named  saman- 
darin.  Faust  describes  samandarin  as  first  stimulating  and  then 
paralyzing  the  automatic  centers  in  the  medulla.  The  poison  resem- 
bles the  alkaloids,  having  the  formula  C2,;Il4„N.O,  and  produces  death 
in  doses  of  0.7-0.9  mg.  per  kilo  (dogs)  with  respirator}^  failure.  Im- 
munization of  rabbits  was  practically  impossible.  A  second  alkaloid, 
samandaridin  (CooH-jiNO)  is  also  present  in  even  greater  quantities 
than  the  samandarin,  and  differs  only  in  being  weaker. 

Fro^s  also  have  similar  poisons  in  their  skins,  extracts  of  Rena  es- 
cidenta  skin  being  highly  toxic."''  The  dermal  secretions  of  most  of 
the  amphibians  are  poisonous,  not  only  for  mammals,  but  also  for  rep- 
tiles, and  in  large  doses  for  the  animals  producing  them  (Phisalix)." 
Bert  '^^  and  also  Dutartre  '^^  have  described  a  digitalis-like  poison  in 
the  secretion  of  the  dermal  glands  of  frogs. 

It  is  evident  that  all  these  poisons  are  quite  distinct  from  the 
venoms,  and  from  the  tn^e  toxins,  apparently  being  simple  chemical 
compounds  not  related  to  the  proteins  and  not  capable  of  causing  im- 
munization. 

POISONOUS  FISH  SI 

There  are  numerous  fish,  especially  in  tropical  waters,  which  de- 
fend themselves  by  injecting  poisons  into  their  enemies.  This  is  ac- 
complished by  spines,  to  which  are  attached  poison  glands.^-  Dun- 
bar-Brunton  ^^  has  described  two  such  fish  (Trachinis  draco  and  Scor- 
pcena  scorpha)  of  Mediterranean  waters.  Wounds  by  these  spines 
cause  in  animals  intense  local  irritation  and  edema  and  paralysis  of 
the  part,  followed  by  gangrene  about  the  site  of  the  wound;  in  fatal 
poisoning  death  occurs  in  from  one  to  sixteen  hours,  with  general  par- 
alysis. The  sufferings  of  persons  so  poisoned  are  said  to  be  extreme, 
and  death  may  occur  either  directly  from  the  poison  or  later  from  sep- 
sis following  the  local  gangrene.  Presumably  this  poison  is  not  dissim- 
ilar to  that  of  the  snakes;  it  probably  is  not  an  alkaloid,  as  Dunbar- 

73a  .Tour.  Pharmacol.,  1910    (S).  .347. 

74Arfh.  pxpor.  Patli.  ii.  l^liarm.,  1S9S    (41).  229    (litoraturo)  :    1900    (43),  84. 

75  Hoppe-Seyier's  Med.  Chein.  T'litrrsucli.,   1801!,  p.  8."i. 

70  Caspar!  and  Loowy,  Med.  Klinik.  1911    (7),   1204. 

77  .Jour.  Phvs.  et  Path,  pen.,  1910   (12),  325. 

78Compt.  Pend.  Soc.  Biol.,   188,5,  p.  524. 

T^Ihid.,  1890,  p.  199. 

81  Full  discussion  and  literature  given  by  Faust.  "Tierische  Gifte."  p.  134. 

82  For  a  list  of  fish  with  poison  {glands  see  Pawlowskv,  Zool.  .Tahrb.,  1912  (.31), 
529. 

83  Lancet,  189G    (ii),  000. 


P0I80N0US  FISH  1^^ 


+o  Tf  nffppts  chiefly  the  heart,  according  to  Pohl,«*  and 
r:r  :ifer,v.i"pSe  w,Lu  l.e,.ave.  UUe  the  ve„..,n  hemo- 
lysins in  that  it  is  acnvated  by  se™^    EvanV  ^^^^^^  ^^  ^^^^^  ^^.^^^ 

to   have    caused   fatal   intoxication   m 

s::^^;^r„ra::."  rz^i^^^^^-  --  - 

iJ  kept  but  ^-/;-^°|^;^_^;^'^:;;:ttle  cannot  be  safely  marketed. 
Tr+n  he  developed  and  contained  in  the  ovaries  and  eggs,  and 

:;  *.,  £r  ...1 1.™.  ~.  aw,,  ..a  —"■•J":"; ;';; 

•„  +i.a  «viim  and  leo-s-  this  terminates  m  collapse,  coma,  ana 
Sr^re^resp-t'oryor  eardiae  paralysis.  Jhe^entire  eo„. 
nf  tl,e  process  may  be  but  ten  to  twenty  minutes,  or  it  may  be  as  many 
lirs  '^On  aeeount  of  the  localization  of  the  poisoii  in  the  eggs  and 
ovaries  not  all  persons  who  eat  the  fish  are  poisoned,  and  not  all  who 
are  poisoneclreeeive  a  fatal  dose.  In  the  gastro-intestinal  form  the 
sWom  appear  later,  consist  chiefly  of  gastro-intestmal  disturbances 
?eZlZ.  more  elosely  ptomain  poisoning,  and  the  prognosis  is  not 

"The  paJhofogilf  ailmy  of  this  foi-m  of  poisoning  has  not  been 
carefully  studied,  but  no  characteristic  or  striking  anatomical  changes 
have  been  noted  in  the  bodies  examined.     Tahara>'  has  described  a 

84Prager  med.  Wocli.,  1S93    (18),  31. 

85  British  Med.  Jour..  1007    (i).73. 

ssaKonstanzoff  and  Manoiloff.  ^^  re.^'^i";      ?   J-';    iSfifi    Hi  ;rature) 

86  "Atlas  des  Poissons  Vencneux,"  St.  Petersburg,  1886    (hteratine). 

STBiochem.  Zeit.,  1010   (30),  2.56. 


164  PnYTOTOXIXS    ASD    ZOOTOXINS 

toxic  body,  tetrodo-toxin,  isolated  from  the  ovaries  of  Tetrodon.^'''^ 
The  purest  preparations  had  a  minimum  lethal  dose  of  0.0025  to  0.004 
grm.  per  kilo,  and  a  provisional  formula  of  CieHgiNOig  was  given  to  it. 
Tetrodotoxin  is  neither  protein  nor  alkaloid,  nor  yet  a  protamin. 

In  tliis  connection  may  be  mentioned  the  peculiar  erysipelas-like 
lesions  caused  by  bites  of  crabs,  which  indicates  the  formation  of  some 
toxic  product  by  tliese  crustaceans.  Gilchrist  **  obtained  a  history 
of  bites  or  injuries  by  crabs  in  323  of  329  cases  of  "  er^-sipeloid. " 
Crabs,  in  turn,  may  be  poisoned  by  cephalopods  which  secrete  an  active 
poison  from  their  salivary  glands.*''  ^lany  coelenterates  produce 
active  poisons  (most  famous  of  these  being  the  Portuguese-man-o'-war), 
which  have  especially  a  paralyzing  and  a  local  irritant  effect.*"'^ 

EEL  SERUM 

In  1888  IMosso  ^°  studied  the  toxic  properties  of  eel  serum,  which  he 
found  was  extremely  poisonous  for  experimental  animals,  0.1  to  0.3  c.c. 
per  kilo  being  fatal  for  rabbits  and  dogs  in  a  few  minutes  if  in- 
travenously injected;  introduced  into  the  stomach  it  is  not  toxic,  but 
it  produces  a  violent  conjunctivitis  when  it  enters  the  eye,  the  poison- 
ous agent  being  contained  in  the  albumin  fraction.^^  The  poisonous 
principle  Mosso  called  ichtJiyoto.rin.  Death  results  from  respiratory 
failure  with  large  doses ;  small  doses  lead  to  cachexia  and  death  after 
a  few  days.  The  coagulability  of  the  blood  is  greatly  reduced.  Kos- 
sel  ®^  found  histological  changes  in  the  central  nervous  system  in  such 
animals,  that  resembled  the  lesions  of  tetanus.  He  succeeded  in  se- 
curing an  active  antitoxin  which  neutralized  the  strongly  hemolytic 
action  of  eel  serum  in  vitro,  and  also  prevented  fatal  effects  in  ani- 
mals. Camus  and  Gley  ^^  have  studied  the  physiological  action  of  eel 
serum  and  found  it  strongly  hemolytic,  and  also  apparently  neuro- 
toxic. The  toxicity  is  destroyed  by  heating"  to  58°  for  fifteen  minutes. 
B}^  immunization  an  antitoxic  serum  can  be  obtained  which  neutralizes 
the  eel  toxin  completely.  Tchistovitch  ^*  secured  antitoxic  serum, 
which  acted  also  as  a  precipitin  for  eel  serum.  De  Lisle  "^  found  that 
eel  serum  does  not  act  like  an  amboceptor,  since  after  heating  it  can- 
not be  reactivated  with  fresh  mammalian  serum,  and  it  seems,  there- 
fore, to  be  different  from  snake  serum  in  its.  structure.  Lamprey 
serum  is  likewise  toxic, ^^  as  is  also  that  of  the  Rays. 

87aAroli.  exp.  Path.  u.  Pharm.,  1890   (26).  401  and  4r).3. 
88  Jour.  Cutaneous  Diseases,  Novemher,  1904. 
soBafrlioni,  Zeit.  f.  T5iol.,   1908    (f)2),  130. 

soa  Roe  von  Fiirth.  Verjrl.  chem.  Phvsiol.;  also  Lojacono,  .Tour.  d.  plivsiol.,  1908 
(10),  1001. 
ooAreli.  Ital.  de  Biol.,  ISSS   (10),  141;   1SS9    (12),  229. 
01  Prdlot  and   Palilson,  Graefe's  Areli.,   1911    (72),   1S:1. 
02Borl.  klin.  Woeh.,   1898    (.35),   ]52. 

03  Arcli.  inU-rnat.  d.  Piiarm..  1899   (.''>),  247. 

04  Ann.  Inst.  Pasteur,   1899    (13),  40fi. 

05  Jour,  of  Mod.  Researoli,   1902   (8).  39fi. 

ooCIev,  ('omi)t.  Rend.  Soe.  Biol.,  1915   (78),  110:  Camus  and  Olev.  ibid.,  p.  203. 


CHAPTER    VII 

CHEMISTRY  OF  THE  IMMUNITY  REACTIONS- 
ANTIGENS,  SPECIFICITY,  ANTITOXINS,  AGGLU- 
TININS, PRECIPITINS,  ANAPHYLAXIS  OR  AL- 
LERGY, ABDERHALDEN  REACTION,  OPSONINS, 
AND  RELATED  SUBJECTS 

Although  immunity  was  first  iuvestigated  in  relation  to  bacterial 
infection,  it  was  soon  learned  that  the  reactions  by  which  the  animal 
body  defends  itself  against  bacteria  have  not  been^.-developed  as 
speeitic  means  of  defense  against  bacteria  alone,  but  are  reactions 
against  foreign  substances  of  similar  chemical  nature,  whether  bac- 
terial, animal,  vegetable  or  artificially  synthetic  in  origin.  Further- 
more, these  reactions  are  chemical  reactions,  and  the  problems  of 
immunity  are  chemical  problems,  although  as  yet  most  of  the  react- 
ing substances  are  not  accessible  to  chemical  investigation.  In  this 
place,  where  our  concern  is  with  the  chemical  aspects  of  pathological 
processes,  the  subject  of  immunity  will  be  discussed  only  from  the 
standpoint  of  the  chemistry  of  the  processes  and  substances  involved, 
leaving  to  other  works  the  clinical  and  bacteriological  aspects  of  the 
subject.^ 

The  reactions  of  immunity  are,  we  find,  reactions  to  chemical  sub- 
stances entering  the  body  from  without,  or  abnormally  developed 
within  the  body  by  invading  organisms  or  by  changes  in  the  chemi- 
cal processes  of  the  body.  Furthermore,  there  seems  to  be  an  essen- 
tial difference  between  the  reactions  incited  by  simple  chemical  com- 
pounds to  which  the  animal  body  can  develop  a  certain  degree  of 
resistance  (such  as  morphine,  alcohol,  and  arsenic),  and  the  reactions 
against  more  complex  substances  such  as  bacterial  toxins,  foreign  pro- 
teins, venoms,  etc.  The  complex  substances  of  the  latter  group  incite 
reactions  wliicli  are  to  a  greater  or  less  degree  specific,  and  usually 
very  highly  augment  the  defense  of  the  body  against  the  foreign  sub- 
stances ;  with  the  simple  poisons  the  reactions  are  largely  or  altogether 
non-specific,  and  the  resulting  resistance  is  usually  relatively  slight. 
Substances  of  the  first  class  we  usually  refer  to  as  antigens. 

1  Especially  to  be  recommended  for  a  discussion  of  the  scientific  problems  of 
immunology  is  Zinsser's  "Infection  and  Resistance,"  Macmillan,  New  York,  1914; 
and  for  methods  and  applications  see  Kolmer's  "'Infection,  Immunity  and  Specific 
Therapy,"  W.  B.  Saunders,  Philadelphia,  1915.  Also  see  Kolle  and*  Wassermann, 
"Handbuch  der  path.  Mikroorganismen";  Weichardt  "Jahresbericht  der  Inimuni- 
tUtsf orschung" ;  and  for  more  recent  literature  consult  the  Zeit.  f.  Immuni- 
tiitsfrsch.,  Referate. 

165 


166  CHEMISTRY    OF    THE    IMMUXITY    REACTIOXS 

ANTIGENS  ■' 

This  term  includes  those  substances  which,  Avhen  introduced  into 
the  blood  or  tissues'  of  an  animal,  in  proper  amounts  and  under 
suitable  conditions,  cause  the  generation  and  appearance  in  the  blood 
of  specific  antibodies  capable  of  reacting  with  the  antigen.  Con- 
cerning the  chemistry  of  antigens  we  can  say  that  all  antigens,  so 
far  as  now  known,  are  colloids.  Furthermore,  with  one  exception, 
every  known  soluble,  complete  protein  may  act  at  least  to  some  degree 
as  an  antigen,  and,  as  yet,  it  has  not  been  finally  established  that  any 
colloids  other  than  proteins  can  act  as  antigens.  The  exception  is 
the  racemized  protein  of  Dakin,  which  Ten  Broeck  -"^  found  to  be  en- 
tirely non-antigenic  although  soluble  and  possessed  of  all  the  amino- 
acids  present  in  the  egg  albumin  used  in  preparing  it.  Solubility  is 
an  essential  character  for  antigenic  action,  for  proteins  that  have  been 
coagulated  by  heat  lose  their  antigenic  capacity,  while  proteins  that 
are  not  coagulated  (e.  g.,  casein,  ovomucoid)  retain  their  antigenic 
properties  after  boiling.-'' 

Of  the  cleavage  products  of  proteins  it  is  certain  that  none  of  the 
amino-acids  and  simple  polypeptids  can  act  as  antigens,  and  it  is 
not  yet  fully  established  that  even  such  large  complexes  as  the  pro- 
teoses are  antigenic,  although  there  is  some  evidence  in  favor  of  this 
view.  AYhether  the  entire  protein  molecule,  or  only  groups  thereof, 
determine  the  characteristics  of  the  antigen,  is  not  known,  there  be- 
ing evidence  w^hich  can  be  interpreted  in  favor  of  either  view,  but 
TVells  and  Osborne  ^  have  submitted  evidence  which  indicates  that  a 
single  protein  molecule  can  act  with  and  engender  more  than  one 
antibody;  this  is  supported  by  Kliein's  demonstration  of  the  produc- 
tion of  two  distinct  antibodies  by  immunizing  with  casein.^'' 

It  has  been  shown  by  Gay  and  Robertson,*  moreover,  that  if  the  non- 
antigenic  cleavage  products  of  casein  are  resynthesized  by  the  re- 
A^erse  action  of  pepsin,  into  a  protein  resembling  paranuclein,  this 
synthetic  protein  is  capable  of  acting  as  an  antigen.  Protamins  and 
globin,  they  found,  were  not  antigenic,^  although  globin  when  com- 
bined with  casein  forms  a  compound  wliicli  engenders  an  antibody  that 
gives  complement  fixation  reactions  witli  globin.  Schmidt  also  found 
that  protomain  edestinate  is  antigenic  for  edestin  and  for  itself,  but 
not  for  protamins,  whereas  a  compound  protein,  both  elements  of 
which  were  non-antigenic  (globin-albumose),  was  not  antigenic. °''- 

2  See  the  Tlevicw  on  Antifrens  by  E.  P.  Pick,  Kolle  ami  Wassorinaiin's  ITaiullnicli 
d.  path.  Mikroorpanismcn,  1912   (1),  G85. 

-2aJour.  Biol.  Chem.,   1914    (17),  369. 
2b  See  Wells,  Jour.  Infect.  Dis.,  1908   (5),  449;  Jour.  Biol.  Clicm..  1910   (28),  11. 

3  Jour.  Infec.  Dis.,  1913    (12),  341. 
-3a  Folia  Microbiol.,   1912    (1),  101. 

,  4  Jour.  Biol,  f'lioni.,  1912   (12),  233. 
■  5  Jour.  Exp.  Mod..  1912   (IG),  479:   1913   (17).  535. 
5a  Univ.  of  Calif.   Puld.,   Pathol..    1910    (2).    l.-i7.     Koviow  and  biblioirrajiliy   on 
specificity. 


NON-PROTEIN  ANTIGENS  167 

NON-PROTEIN  ANTIGENS 

Amono-  the  many  aeeouiits  of  what  the  autliors  interpret  as  the 
successful  production  of  specific  antibodies  as  a  reaction  to  non-pro- 
tein antigens,  are  the  following: 

Ford  **  found  that  rabbits  can  be  immunized  to  extracts  of  Aman- 
ita phalloides,  and  that  the  serum  of  such  rabbits  will  neutralize  five 
to  ei^-lit  times  the  lethal  dose  for  guinea-pi«>'s.  and  is  anti-hemolytic 
for  tlie  hemolysin  of  amanita  when  diluted  to  1-1000.  As  he  and 
Abel "  had  found  this  hemolytic  poison  of  Amanita  to  be  a  glucoside, 
this  observation  is  to  be  interpreted  as  a  successful  production  of  an 
antibody  for  a  non-protein  poison,  a  glucoside.  This  work  was  fur- 
ther 'supported  by  successfully  immunizing  rabbits  to  extracts  of 
Rhiis  toxicodendron,  and  finding  that  their  serum  in  doses  of  1  cc. 
will  protect  guinea-pigs  from  5-6  lethal  doses  of  the  poison,  which 
was  found  by  Acree  and  Syme  ^  to  be  a  glucoside.  Subsequent  work 
by  the  same  author  confirms  the  main  point,  showing  that  an  active 
hemolysin  can  be  obtained  free  from  demonstrable  protein,  and  that 
immunization  with  this  protein-free  hemolj'sin  will  result  in  stronglj' 
active  (1-1000)  antihemolytic  serum.^ 

Ajiother.  non-hemolytic  poison  from  Amanita,  which  Ford  desig- 
nates as  Amanita  toxin,  was  found  to  contain  neither  protein  nor 
glucoside,  and  no  antitoxic  serum  or  definite  artificial  immunity 
can  be  obtained  for  it.  The  antihemolysin  unites  with  the  hemoly- 
sin in  simple  multiple  proportions. ^° 

Jaeoby  believed  that  he  had  obtained  the  phytotoxin  ricin  free  from 
protein,  in  which  case  the  well-known  and  active  antiricin  must  rep- 
resent an  antibody  for  a  non-protein  antigen.  However,  the  work 
of  Osborne,  iMendel  and  Harris  ^^  has  shown  that  ricin  is,  in  all 
probability,  an  albumin,  and  this,  for  the  present  at  least,  places  ricin 
with  the  protein  antigens. 

The  work  of  Ford  is,  in  our  estimation,  the  strongest  evidence  yet 
presented  as  to  the  possibility  of  non-protein  antigens.  The  newer 
developments  in  immunological  research,  moreover,  make  it  seem 
entirely  plausible  that  a  complex  glucoside,  which  can  be  hydroh'zed 
by  enzymes,  can  act  as  an  antigen.  If  we  consider  the  evidence  that 
immunity  consists  in  the  development  of  a  special  power  to  hydro- 
lyze  foreign  substances,  when  these  substances  are  of  such  a  nature 
as  to  stimulate  the  cells  to  activity,  and  that  Abderhalden  and  others 
have  found  evidence  that  specific  enzymatic  properties  appear  in 
the  blood  of  animals  injected  with  carbohydrates  and  fats,  it  seems 

c.Tour.  Tnfec.  Dis.,  1907  (4),  541. 
T  Jour.  Biol.  Chem.,  1907  (2),  273. 
8. Jour.   Biol.   Chem..    1007    (2),   547. 

9  Jour.  Pharmacol..  1910   (2),  145. 

10  Jour.  Pharmacol.,  191.3    (4),  235. 

11  Amer.  Jour.  Physiol.,  1905    (14),  259. 


168  CHEMISTRY    OF    THE    IMMUNITY    REACTIONS 

entirely  reasonable  that  a  toxic  glucoside  can  have  antigenic  proper- 
ties. A  similar  line  of  reasoning  will  apply  to  the  question  of  lipoid 
antigens. 

The  evident  participation  of  lipoids  ^-  in  innnunity  reactions,  es- 
pecially the  complement-fixation  and  allied  reactions,  has  naturally 
led  to  investigation  of  the  possibility  that  lipoids  may  act  as  true 
antigens,  a  possibility^  made  conspicuous  by  the  fact  that  lipoids  can 
be  substituted  for  true  antigens  in  the  "Wassermann  reaction  (q.  v.). 
Bang  and  Forssmann  immunized  with  ethereal  extracts  of  red  cor- 
puscles and  obtained  hemolysins,  so  that  they  concluded  that  the 
antigenic  constituent  of  the  corpuscles  is  a  lipoid,  probably  a  phos- 
phatid.  This  Avork  has  caused  much  controversy  and  many  workers 
have  failed  to  confirm  their  results.^^  It  is  a  striking  fact  that  when 
purified  phosphatids,  from  sources  favorable  for  obtaining  pure  ma- 
terials, are  used,  the  results  are  always  negative,  while  the  positive 
results  are  generally  reported  with  lipoids  of  more  or  less  dubious 
purity. 

IMuch  and  others  have  worked  with  lipoids  from  a  streptothrix, 
which  is  called  "nastin,"  and  they  state  that  sera  are  obtained 
which  give  complement  fixation  reactions  with  nastin  used  as  the 
antigen.^*  Similar  results  are  described  for  the  fatty  materials  from 
tubercle  bacilli  ( ' '  tuberculonastin  " ) . 

Meyers  ^^  has  reported  the  production  of  specific  complement  fix- 
ation antibodies  by  immunizing  rabbits  with  acetone-insoluble 
lipoidal  material  obtained  from  tape  worms  and  echinococcus.  He 
has  found  the  acetone-insoluble  fraction  of  tubercle  bacilli,  presum- 
ably phosphatids,  to  serve  as  antigen  in  complement  fixation  reac- 
tions with  antibodies  for  tubercle  bacilli,^*'  and  much  more  effectively 
than  the  protein  residue  of  the  bacilli,  wherefore  he  concludes 
that  the  reactions  obtained  with  the  lipoids  certainly  cannot  be 
ascribed  to  adherent  traces  of  protein.  Bergel "  observed  after 
lecithin  injections  in  rabbits,  not  only  an  increase  in  the  lipase  con- 
tent of  the  blood  and  tissues,  but  also  the  presence  of  complement- 
binding  antibodies,  and  -Tobling  and  Bull  ^^  have  found  an  increase  in 
-:erum  li])ase  after  imiimnizing  with  red  corpuscles. 

Bogomolez  ^"  suggests  that  the  lipoids  themselves  may  be  produced 
in  excess  for  defense  against  various  poisons,  which  they  serve  to 
inhibit,  especially  the  toxin  of  B.  hotulinus. 

12  Bibliography  on  Lipoids  and  Immunity  pivon  by  Landsteincr.  KoUo  and  Was- 
sormann's  TTandbnoh,  101.3    (2),  1240;  .Toblino;,  Jour.  Tmmnnol.,   1010    (1).  401. 

13  Review  of  literature  bv  T.andsteiner.  Jaliresb.  Imnninitiitsfrscli..  1010  (fi),. 
200. 

14  Literature  in   Beitr.  Klinik  d.  Tuberlc,  1011    f20).  341. 
isZeit.  Immunitiit.,  1010   (7),  7.32;   1011    (9),  530;   1012    (14),  355. 
if>  Ihid.,   1012    (14),  350;    1012    (15).  245. 

i7Deut.  Areh.  klin.  Med..  1012    (106),  47. 

15  Jour.  Kxp.  Med.,  1012   (10),  4S3. 
i«Zeit.  Immunitiit.,  1010    (S),  35. 


yoS-rilOTEIS  ASTKIESH  169 

The  number  of  reputed  positive  results  witli  lipoids  makes  it  im- 
possible at  this  time  to  state  dogmatically  that  lipoids  may  not  pos- 
sess antio-enic  propcM-ties,  but  it  must  be  taken  into  account  that  the 
successful  use  of  lipoids  as  antigens  in  complement  fixation  reactions 
is  not  proof  of  their  true  antigenic  nature,  in  view  of  our  present  lack 
(,f  knowledge  of  the  actual  nature  of  this  reaction  itself,     i^  urther- 
more    we  have  the  testimony  of  Fitzgerald   and  Leathes  ^«   that  a 
lipoidal  material  from  liver,  which  was  itself  capable  of  serving  as 
antio-en  in  the  Wassermann  reaction,  did  not  engender  complement- 
fixiiro-  antibodies  in  rabbits  immunized  with  this  lipoid.     Kitchie  and 
Mille"r==^  could  find  no  antigenic  activity  in  the  lipoids  of  serum  or 
corpuscles.     Also  Kleinschmidt,^^   who  accepts  the  antigenic  nature 
of  nastin    was  unable  to   secure   antibodies   by   immunizing  rabbits 
with  nastin.     Thiele  =^^  calls  attention  to  the  fact  that  lipoids  possess 
no  specificity,  and  therefore  cannot  give  rise  to  antibodies.     Neufeld 
found  that  "rabbits  immunized  with  lecithin  developed  no  opsonins 
for  lecitliin  emulsions.     A  suggestive  observation  is  that  of  Pick  and 
Schwarz  -*  who  found  that  the  presence  of  lecithin  increases  the  anti- 
genic power  of  bacteria,  which  may  help  to  explain  the  activity  of 
possible  traces  of  proteins  in  lipoid  preparations  used  as  antigens. 

Many  drugs  cause  a  hypersensitization,  and  in  this  respect  seem  to 
behave  as  antigens  produciirg  anaphylactic  antibodies.  It  happens 
that  most  of  these  chemicals  are  of  such  a  nature  as  to  permit  of  their 
nnion  with  proteins,  and  it  seems  probable  that  such  protein  com- 
pounds behave  as  foreign  proteins  to  the  animal  in  which  they  are 
formed,  for  it  has  been  found  that  guinea-pig  seram  treated  with 
iodin  can  render  guinea-pigs  sensitive  to  the  same  iodized  serum.- 
Hence,  hypersensitiveness  to  iodin  compounds  would  be  a  reaction  to 
iodized  proteins,-"  and  not  to  the  non-protein  iodin  compound:  the 
same  applies  to  anaphylactic  reactions  observed  with  salvarsan, 
atoxyl,"  and  perhaps  aspirin  and  antipyrin.=«  Zieler,  however,  has 
questioned  the  validity  of  many  of  the  experiments  on  which  these 
views  are  based.^«  It  is  possible  that  certain  chemicals  may  react  m 
such  a  way  with  the  tissue  or  blood  proteins  as  to  make  them  sensi- 
tive to  the  animal's  own  complement,  which  then  forms  anaphyla- 

20  Univ.  of  Calif.  Publ.,  1912  (2) ,  39. 

2iJoiir.  Path,  and  Bact.,  1013   (17),  429. 

22Berl.  klin.  Woch.,  1910  (47),  57. 

23Zeit.  Imnmnitat..   1913    (IG),   160. 

24Biochem.  Zeit..  1909   (15).  4.53. 

25  Friedborger  and  Ito,  Zeit.  Immunitiit.,  1912    (12),  241. 

2GAccordincr  to  Block    (Zeit.  exp.  Path.,  1911    (9).  509)    iodoform  uhosyncrasv 
depends  upon  the  CH.  rather  than  on  the  iodin,  and  is  a  local  cellular  reaction 
rather  than  a  humoral  reaction,  the  protoplasm  havm-  an  moroased  affinity  for 
methyl  radicals.      (See  Weil.  Zeit.  Chemotherapie    1913    (1-412.) 
_^  27Moro  and  Stheeman,  Miinch.  med.  Woch.,  1909    (56),  1414.  ,     „.     , 

-    28Bruck,  Berl.  klin.   Woch.,   1910    (47),    1928;    Klausner,  Miinch.   med.  ^^  och., 
1911    (5S),   138. 

29  Munch,  med.  Woch.,  1912   (59),  401. 


170  CHEMISTRY    OF    THE    IMMUXITY    REACTIOXS 

toxin,^°  and  thus  causes  reactions,  but  the  whole  anaphylatoxin  ques- 
tion is  in  so  uncertain  a  state  at  the  time  of  writing  that  further 
speculation  in  this  direction  is  not  justifiable. 

The  attempts  to  produce  antitoxin  against  cantharidin  have  not 
yielded  convincing  results,-'^  nor  against  epinephrine.^-  De  Angelis  ^^ 
claimed  that  he  had  produced  specific  precipitins  for  various  natural 
and  syntlietic  dyes,  but  this  woi'k  has,  as  was  to  be  expected,  failed 
of  confirmation.^^  Elschnig  and  Salus ""  state  that  melanin  from  the 
eye  is  antigenic,  producing  complement-fijiing  antibodies  specific  for 
melanin  but  not  for  the  species.  We  know  too  little  concerning  the 
composition  of  melanin  to  interpret  this  observation;  furthermore, 
their  preparation  was  not  tested  for  proteins. 

In  general  terms,  therefore,  antigens  are  protein  molecules,  and 
the  reactions  of  immunity  are  reactions  against  proteins  foreign  to 
the  body  of  the  host,  and  manifested  by  the  presence  in  the  blood 
of  the  reacting  animal  of  substances  which  combine  with  and  cause 
recognizable  changes  in  the  foreign  protein.'"  These  changes  are 
recognized  in  many  ways,  such  as  precipitation,  agglutination,  com- 
plement-fixation, etc.,  and  the  question  at  once  arises  as  to  whether 
these  diiferent  manifestations  depend  each  upon  a  separate  antibody, 
or  if  several  or  all  of  them  are  not  caused  by  a  single  antibod}^  the 
action  of  which  is  indicated  by  the  different  reactions  which  are  made 
manifest  by  different  procedures  in  each  case.  This  question  will  be 
discussed  further  in  later  paragraplis. 

Knowing  that  the  antigens  are  merely  foreign  proteins  which  have 
been  introduced  into  the  body  of  an  animal,  there  naturally  occurs 
the  thought  that  the  animal  body  is  continually  receiving  in  its  food 
foreign  proteins,  and  against  which  it  defends  itself  in  the  alimentary 
canal  by  enzymatic  action,  which  disintegrates  these  proteins  until 
they  have  lost  their  colloidal  character.  Logically  following  this 
comes  the  idea  that  perhaps  the  reactions  of  immunity  are  simply 
the  same  or  similar  disintegrative  enzymatic  actions,  carried  on  within 
the  blood  and  tissues  to  protect  the  body  in  the  same  way  against 

30  See  Manoilov,  Wien.  klin.  Woch.,  1912   (25),  1701. 

31  Champy,  Compt.  Rend.  Soc.  Biol.,  Ifl07    (62),  1128. 
32Pollak,  Zeit.  pliysiol.  Clieni.,   1010    (68),  GO. 

33  Ann.  di  Ig.  Sperim.,  1009    (19),  33. 

34Takemura.  Zoit.  Imnuinitiit.,  1010   (5),  607. 
__  35Graefe's  Arch.,  1011    (70),  428. 

3C  Drew  lias  found  no  evidence  of  antibody  formation  Ity  imnuinii'inti  molluscs 
and  ecliinoderms  (Jour,  of  Hyp.,  1011  (11),  188),  from  which  ho  conchules  that 
the  reaction  to  foreign  pioteins  is  not  a  imiversal  ]iroperty  of  pmloplasm ;  a 
sweeping  generalization  which  requires  more  extensive  investigation  for  its  es- 
tablisliiiiciil.  ("antacuzenp  (('omi)t.  Rend.  Soc.  Biol.,  1013  (74).  Ill)  obtained 
precipitins  l)y  iiniiiuni/.ing  J'linlliisia  itiiimiJlatd  witii  mammalian  blood,  but  no 
licmolysins  with  this  or  A plirodilr  aciilcala  and  Elcdonc  vioschatn.  Carrel  and 
Ingebrigtsen  (.Tour.  Kxp.  ]\Ied.,  1012  (l.'i),  287)  have  found  that  tissues  growing 
iti  vitro  with  foreign  blood  jjroduce  hemolytic  antibodies  for  that  blood,  indicat- 
ing that  isolated  cells  can  react  to  antigens  by  antiliody  ]>niduction. 


SPECIFICITY  OF  IMMVyF  h'FACTIOXS  171 

foreign  proteins  whicli  tiic  alimentary  digestive  apparatus  has  not 
had  the  opportunity  to  destroy.  This  conception  of  the  nature  of  im- 
mune reactions  to  antigens  has  been  especially  advanced  and  in- 
vestigated by  Victor  C.  Vaughan  ^^"^  and  his  co-workers,  and  l)y  Kmil 
Abderhalden,  who  has  demonstrated  in  various  ways  an  increased 
proteolytic  power  in  the  blood  of  animals  which  have  received  pa- 
renteral injections  of  foreign  proteins.'"''  Thus,  if  the  antiserum  re- 
acts on  the  specific  protein  within  a  dialyzing  sac,  the  products  of 
proteolysis  diffuse  into  the  surrounding  medium,  where  they  can 
be  detected  by  simple  chemical  reactions.  Also,  changes  in  the  spe- 
cific rotation  of  the  protein  or  peptid  solution  can  be  observed  by 
the  polariscopic  reading  before  and  after  the  action  of  the  antiserum. 
A  particularly  important  corroboration  of  Vaughan 's  theory  is  fur- 
nished by  tlie  behavior  of  the  racemized  i)rotein  of  Dakin.  Although 
soluble,  this  protein  cannot  be  attacked  by  the  digestive  proteolytic 
enzymes,  presumably  because  of  its  altered  configuration ;  and  it"  is 
non-antigenic,  presumably  because  it  cannot  be  attacked  by  the  pro- 
teases of  the  blood  and  tissues.  Likewise  it  cannot  be  nietabolized, 
w^hether  fed  or  injected  subcutaneously.^"^  Here  we  have  good  evi- 
dence of  the  fundamental  identity  of  the  three  processes,  digestion, 
metabolism,  antigenic  activity. 

As  immunity  reactions  manifest  themselves,  however,  there  are  many 
steps  in  the  process  besides  simple  hydrolysis  of  proteins,  even  if 
This  be  the  ultimate  goal  of  them  all. 

SPECIFICITY  OF  IMMUNE  REACTIONS 

The  many  attempts  to  explain  the  various  reactions  of  immunity 
solely  on  the  basis  of  known  physico-chemical  properties  of  colloids 
all  flatten  out  when  the  striking,  characteristic,  and  often  extreme 
specificity  of  these  reactions  is  considered.  Chemical  explanations 
are  but  little  more  satisfactory.  In  enzyme  action  we  find  many  com- 
parable examples  of  specificity, — but  this  does  not  help,  as  the  enzymes 
are  as  mysterious  as  the  antibodies.  But  no  proposed  explanation  of 
any  of  the  reactions  incited  by  antigens  can  be  of  value  if  it  fails  to 
take  into  account  the  specificity  of  the  reactions.  We  lack  the  space 
here  to  consider  the  many  ideas  and  the  items  of  evidence  which  have 
been  advanced  concerning  this  all-important  chemical  problem,  but 
refer  the  reader  to  the  excellent  discussion  by  E.  P.  Pick-^^"*  The  main 
facts  at  present  available  are  the  following:  Specificity  was  at  first 
supposed  to  depend  solely  upon  biological  relationships,  for  it  was 
found  easy  to  distinguish  the  serum  of  animals  of  unlike  nature  by 
n  cans  of  the  precipitin  and  other  reactions,  but  the  more  closely  re- 

-6a  See  Vaiiglmn,  "Protein  Split  Products,"  Pliiladelpliia,  1913. 
3"  Abderhalden,  "Abwehrferniento  dos  tierisclicn  Oriranismus,"  Perlin.   10]  3. 
"a  See  Ten  Broeck,  Jour.  Biol.  Cliem.,  1014    (17).  3(1!). 

3S  Kolle  and  Wassermann's  Ilandhuch  d.  path.  ^Mikroor^ranismen,  1012  (1), 
685 ;   full  bibliography. 


172  CHEMISTRY    OF    THE    IMMVSITY    REACTIONS 

lated  the  animals  the  less  sharply  these  reactions  distinguish  them, 
until,  with  such  closely  related  animals  as  dog  and  fox,  or  man  and 
apes,  antisera  for  the  blood  of  one  react  nearly  as  well  with  the 
blood  of  the  other,  the  existing  differences  being  only  quantitative. 
The  opinion  therefore  gained  ground  that  the  specificity  depended 
upon  some  peculiar  biological  relationship  of  the  antigens,  and,  as 
serum  proteins  which  seemed  to  be  quite  similar  chemically  but  which 
were  from  unrelated  species,  were  sharply  differentiated  by  the  bio- 
logical reactions,  that  the  specificity  must  depend  upon  something 
quite  distinct  from  ordinary  chemical  differences.  But  even  with 
closely  related  species,  differences  can  often  be  brought  out  by  means 
of  the  process  of  saturation  (which  consists  in  treating  the  antiserum 
with  sufficient  quantities  of  an  antigen  until  it  no  longer  reacts  with 
additional  quantities  of  this  antigen,  and  then  trying  its  reactive 
power  with  the  other  related  antigen  which  one  wishes  to  test). 

As  use  began  to  be  made  of  other  materials  than  serum,  and 
especially  when  more  or  less  purified  proteins  were  employed,  it  was 
found  that  within  the  tissues  of  a  single  animal  or  plant  there  might 
exist  antigens  which  were  quite  distinct  from  one  another — more  so, 
indeed,  than  some  of  the  chemically  similar  substances  of  different 
biological  origins.  Thus,  in  the  hen's  egg,  by  means  of  the  anaph}'- 
laxis  reaction,  I  have  been  able  to  distinguish  five  distinct  antigens, 
and  these  correspond  to  as  many  different  proteins  which  have  been 
distinguished  by  chemical  means. ^''  Also,  for  another  example,  in 
the  crystalline  lens  are  found  proteins  which  are  specific  for  lens 
proteins,  in  that  they  produce  antibodies  reacting  with  lens  proteins 
from  varied  species  of  animals,  but  not  with  the  serum  proteins  of 
the  species  from  which  the  antigenic  lens  substance  was  derived.*" 
Here  the  chemical  character  of  the  protein  is  undoubtedly  more 
^significant  than  its  biological  relations.  These  and  other  observa- 
tions leave  little  room  for  doubt  that  specificity  does  depend  upon 
chemical  composition,  and  that  the  differences  in  species  as  exhibited 
hy  their  biological  reactions  depend  iipon  distinct  differences  in  the 
chemistry  of  their  proteins.*°'^  Chemically  distinct  proteins  {e.  g.  lens 
and  serum  proteins)  of  one  animal  may  be  immunologically  distinct, 
and  chemically  related  proteins  of  dissimilar  species  (c.  g.  casein 
from  goat  and  cow  milk)  may  shoAV  immunological  relationship. 
Crystalline  albumin  from  hen's  eggs  shows  no  immunological  distinc- 
tion fi-om  that  of  ducks'  eggs,  whereas  eacli  of  the  three  ju'oteins  sep- 
arable fi'om  horse  serum — euglobulin,  pseudoglobulin  and  albumin — 
can  l)e  distinguished  from  the  other  two  l)y  the  anaphylaxis  reac- 
tion.''"''    Furthermore,   it   h.as   been  shown   by   AVells   and   Osborne  *^ 

30  Jour.  Infeo.  Dis.,  1911   (9),  147. 

40  Spc  Knisiua,  Zcit.  Immunitiit.,  1910   (5),  tlOi). 

40ii8ee  WoUs  and  Oaboriic,  Jour.  Infect.  Dis..  1910   (19),  183. 

•if'hDalc  and  Ilardov,  Hiocliem.  Jour.,  191G   (10),  408. 

■ii.ldwr.    Infect.  Dis.",   191.3    (12),  341. 


si'Kcu'JciT)  or  ni]ii  \K  /.'/v.kwo.v.s'  173 

that  a  single  pure  protein  may  exliibit  jiiultiple  antigenic  properties, 
and  react  or  fail  to  react  with  other  pure  proteins  according  to 
whether  chemical  differences  can  be  demonstrated  by  recognized 
analytical  methods. 

A  striking  example  of  the  existence  of  identical  antigenic  prop- 
erties in  materials  of  biologically  unrelated  origins,  is  furnished  by 
the  sheep  corpuscle  hemolysin  discovered  by  Forssner,*^-''  who  found 
that  many  different  materials,  when  injected  into  rabbits,  engender  in 
the  rabbits'  serum  a(!tive  hemolytic  amboceptors  for  sheep  corpuscles. 
This  antigenic  property  has  been  demonstrated  in  the  organs  of  the 
guinea-i)ig,  horse,  cat,  dog,  mouse,  cliicken,  turtle,  and  several  species 
of  fish,*^^  although  not  exhibited  by  organs  of  many  closely  related 
species  (>.  g.  pig,  ox,  rabbit,  goose,  frog,  eel,  man,  pigeon,  rat).  It  is 
not  present  in  the  red  corpuscles  of  these  animals,  but  is  present  in 
the  corpuscles  of  the  sheep,  whose  organs  do  not  have  this  property. 
It  has  also  been  found  in  paratyphoid  and  Gartner  bacilli,  mouse 
tumors  and  sheep  spermatozoa.  Not  only  does  the  serum  of  rabbits 
thus  inmiunized  show  active  hemolysis  for  sheep  corpuscles,  but  if 
injected  into  the  vein  of  an  animal  whose  organs  contain  this  antigen 
there  results  a  prompt,  severe  anaphylactic  intoxication,  presumably 
Through  reaction  between  the  antigen  present  in  their  tissues  and  the 
antibodies  of  the  rabbit  serum.  Furthermore,  the  antibody  can  be 
specitically  removed  from  the  immune  rabbit  serum  by  contact  with 
any  of  the  antigen-containing  tissues,  but  not  by  tissues  that  do  not 
exhibit  this  antigenic  property.  The  antigen  seems  to  remain  in  the 
tissues  when  the  fluids  are  forced  out  by  pressure,  and  Doerr  and 
Pick  believe  it  to  be  associated  with  the  nucleoproteins. 

This  series  of  observations,  which  seem  to  have  been  quite  gen- 
erally corroborated,  indicates  conclusively  that  the  immunological 
specificity  of  an  antigen  is  not  necessarily  related  to  the  biological 
specificity  of  the  living  organism  from  which  it  is  derived.  The  logi- 
cal explanation  is  that  there  ma.v  exist  proteins  in  different  species 
which  have  chemical  resemblances  or  identity,  and  this  is  scarcely  to 
be  doubted.  We  find  identical  lipoids,  fats,  nucleic  acids,  and  carbo- 
hydrates in  different  species ;  many  peculiar  types  of  proteins  show 
apparent  chemical  identity  in  different  species  (e.  g.  gelatin,  keratin)  ; 
some  chemically  similar,  derived  proteins  also  seem  immunologically 
identical  or  closely  related  (e.  g.  lens  protein,  casein).  Therefore,  it 
is  highly  probable  that  many  tissue  proteins  may  be  identical  in  dif- 
ferent forms  of  animal  cells,  and  even  in  animal  and  plant  cells. 

Another  sort  of  manifestation  of  apparently  non-specific  immunity 
reactions  has  been  observed  especially  in  therapeutic  immunizations.*^'' 
Beginning  with  the  classical  observation  of  Matthes  that  the  tuber- 

4ia  Review  by  Doerr  and  Pick,  Rioeliem.  Zeit..  1014    (00),  2r)T. 

4ibTsunecka*  Zeit.  Inimiinitat.,   1014    (22),  ,567. 

4ic  See  review  by  Jobling,  .Tour.  Amer.  Med.  Assoc.,  1916    (66),  1753. 


174  CHEMISTRY    OF    THE    IMMUNITY    REACTIOXS 

ciiliii  reaction  could  be  produced  with  deutero-albumose,  many  sim- 
ilar non-specific  reactions  have  been  observed.  Particularly  the 
sharp  reaction  that  follows  intravenous  injections  of  killed  typhoid 
bacilli  into  typhoid  patients  has  been  found  to  result  equally  well  if 
colon  bacilli  are  used,  or  deutero-albumose.  One  possible  explana- 
tion of  this  type  of  reaction  is  that  the  injected  substance  acts  as  a 
common  antigen,  which  causes  the  production  of  common  antibodies 
that  react  also  with  the  antigens  of  the  cause  of  the  disease.  Another 
possibility  is  that  the  foreign  protein  stimulates  the  tissues  that 
form  antibodies,  presumabl.y  the  red  marrow,  so  that  they  produce 
not  only  antibodies  for  this  antigen,  but  also  for  the  antigens  of  the 
specific  etiologic  factor  of  the  disease  that  have  been  stimulating  the 
bone  marrow  previously.  INIoreover,  the  febrile  reaction,  the  leu- 
cocytosis,  and  other  phenomena,  such  as  the  antiferment  index  of 
the  serum  ( Jobling),  that  injection  of  nonspecific  protein  produces, 
may  be  responsible  for  favorably  affecting  the  disease,  rather  than 
actual  antibody  formation. 

An  interesting  illustration  of  the  fact  that  whatever  stimulates  the 
bone  marrow  may  cause  it  to  form,  among  other  blood  elements,  spe- 
cific antibodies,  is  furnished  by  the  behavior  of  antitoxin-producing 
horses.  If  a  horse  that  has  been  imnuinized  to  diphtheria  toxin  is 
bled  as  much  as  possible,  it  will  be  found  to  have  regenerated  the  lost 
antitoxin  within  48  hours,*^*^  even  although  the  last  immunizing  dose 
of  toxin  was  received  long  before.  Also,  it  is  stated  that  persons  who 
have  once  had  typhoid,  but  whose  blood  no  longer  contains  much 
agglutinin,  may  show  a  high  typhoid  agglutinin  content  when  infected 
by  some  other  organism,  or  after  any  sharp  febrile  attack.  It  is 
highly  possible  that  many  therapeutic  agents  may  similarly  act  by 
stimulating  the  marrow  to  increased  formation  of  specific  antibodies, 
e.  g.  arsenic,  mercury  and  other  metals,  lieliotlierapy,  hemorrhage  or 
phlel)otom.y,  hot  baths. 

The  other  aspect  of  specificity,  i.  e.,  the  presence  of  several  antigens 
in  a  single  organism,  entirely  distinct  from  other  antigeus  in  the  same 
organism,  has  been  repeatedly  demonstrated.  Besides  the  identifica- 
tion of  five  distinct  antigens  in  tlie  hen's  ogfc.  mentioned  previously, 
we  have  the  repeatedly  demonstrated  individuality  of  serum  pro- 
teins and  milk  casein  of  the  same  animal,  and  even  the  differentiation 
of  casein  from  lactalbumin  in  the  same  milk,  as  contrasted  witli  the 
common  inter-reactions  of  caseins  from  different  sources,^^®  e.  g.,  cow 
and  goat.  A  certain  but  slight  distinguishable  specificity  may  be  ob- 
served between  proteins  from  different  organs  of  the  same  animal, 
which  differentiation  is  still  sharper  between  the  tissue  proteins  and 
serum   proteins   of    tlic    }iniiii;i].^"'     Sex    cells    esjiecially    seiMu    to    be 

iidO'Rripn.  .Jour.  Palli.  iiiul   Had.,  ini.3    (18),  SO. 
4ic.Sce  VorscU.  Zeit.  liiiiimnitiit.,  1915   (24),  2()7. 
4ifSee  Salus,  liiocliein.  Zeit.,    ION    (GO),   1. 


SPECIFICITY  OF  IMMUNE  REACTIONS  175 

rather  distinct  iminnnol()<iically  from  the  body  cells. ■'^°  Numerous 
instances  of  two  separate  proteins  from  the  same  plant  seeds  show- 
ing entirely  distinct  immunological  specificities  have  been  de- 
scribed/^'* On  the  other  hand,  hemoglobins  which  the  crystallo- 
graphic  work  of  Reichert  has  shown  to  be  remarka])ly  specific  from 
that  standpoint,  seem  to  show  an  equally  marked  si)ecies  specificity 
when  tested  by  anaphylaxis.^^'  Indeed,  some  of  the  most  striking 
examples  of  absolute  specificity  are  furnished  by  red  corpuscles,  which 
show  readily  demonstrable  differences  between  closely  related  indi- 
viduals. For  example,  take  the  remarkable  observation  of  Todd,*^^ 
who  mixed  together  isolytic  beef  sera  from  over  60  animals,  and  then 
tested  the  mixture  with  the  corpuscles  of  110  different  cattle,  all  of 
which  were  hemolyzed.  But  when  the  mixture  of  sera  was  ex- 
hausted with  the  corpuscles  of  any  one  of  the  110  cattle  it  would 
then  hemolyze  the  corpuscles  of  all  the  other  109,  but  was  absolutely 
without  action  on  the  corpuscles  of  the  individual  with  whose  cor- 
puscles it  had  been  exhausted.  This  indicates  that  the  red  corpuscles 
of  any  individual  possess  characters  which  differentiate  them  from 
the  corpuscles  of  any  other  individual  even  of  the  same  species. 

As  satisfactory  a  conception  of  the  nature  of  specificity  as  our 
present  evidence  warrants  is  that  developed  by  Pick,  largely  on  the 
basis  of  his  own  work.  He  properly  accepts  the  influence  of  both 
the  physico-chemical  properties  and  the  chemical  composition  of  the 
colloids  concerned  in  immunity  reactions  as  determining  specificity. 
Both  these  factors  undoubtedly  come  into  play  in  determining  the 
possibility  of  interaction  of  antigen  and  antibody.  The  electric 
charges  of  the  amphoteric  colloidal  antigen  and  antibody,  and  per- 
haps also  their  surface  configuration  and  their  surface  forces,  all 
influence  their  reaction ;  these  physico-chemical  factors  greatly  com- 
plicate the  possibility  of  reaction  between  two  colloids,  and  to  these 
influences  are  added  the  influence  of  the  chemical  structure  in  deter- 
mining subsequent  chemical  reactions.  It  would  seem  possible  that 
the  existence  of  all  these  factors  may  account  for  specificity,  it  being 
necessary  for  each  one  of  a  long  series  of  both  physical  and  chemical 
adjustments  to  agree  perfectly  in  order  that  reaction  may  take  place 
— just  as  in  a  combination  lock  one  lever  after  another  is  thrown  by 
the  i)roper  manipulation  of  the  dial,  and  only  when  all  the  long  series 
of  levers  is  in  just  the  proper  position  does  the  bolt  engage  and  the 
lock  open." 

The  studies  of  Pick  and  his  colleagues,  amplified  somewhat  by  other 
investigations,  have  led  to  the  following  view  of  the  chemistry  of 

4igGraetz,  Zeit.  Immunitiit.,  1014   (21),  150. 

4ih  Wells  and  Osborne,  Jour.  Infect.  Dis.,  1911    (8),  GG  et  sen.,  especially  lOlG 
(19),  18.3. 
4ii  Bradley  and  Sansum,  Jour.  Biol.  Cliem.,  1914    (18),  407. 
4ij,Jour.  of  Genetics,  1913   (3),  123. 
42  The  "resonance  tlieory"  of  Traube  assumes  tliat  the  surface  forces  of  react- 


176  CHEMIHTRY    OF    THE    IMMJ-XITY    REACTIOXS 

specificity:  There  exist  two  sorts  of  specificity  in  each  protein 
molecule;  one  of  these  is  easily  altered  by  simple  physical  measures, 
e.  g.,  heat,  cold,  partial  coagulation,  etc.,  without  essentially  chang- 
ing the  chemical  composition  of  the  protein.  AVheu  so  altered  the 
antigenic  properties  of  the  protein  are  likewise  altered,  in  that  the 
antibody  it  engenders  differs  somewhat  in  the  scope  of  its  reactivity, 
from  the  antibody  engendered  by  the  original  unaltered  protein;  but 
the  alteration  does  not  affect  the  species  characteristics  of  the  antigen. 
Thus,  a  heated  antigen  may  engender  ]-)recipitins  that  will  react  with 
this  heated  antigen,  but  not  with  similar  heated  proteins  from  other 
species  of  animals,  while  the  antibodies  engendered  by  the  same  but 
unheated  antigen  will  not  react  with  the  heated  protein. 

The  other  sort  of  specificity  is  not  so  easily  affected,  only  marked 
chemical  alterations  of  the  antigen  modifying  it,  and  this  concerns  the 
species  characteristics  of  the  antigen.  This  fundamental  species  speci- 
ficity seems  to  be  closely  related  to  the  aromatic  radicals  of  the  protein 
antigen,  for  it  is  affected  by  introducing  into  the  protein  molecules 
substances  which  are  known  to  combine  with  the  benzene  ring, 
e.  g.,  iodin,  diazo  and  nitro  groups.  Proteins  thus  chemically  altered 
will  act  as  proteins  foreign  to  animals  of  the  species  from  which  they 
are  derived,  and  the  antigens  they  develop  are  devoid  of  species 
specificity,  although  quite  specific  for  proteins  like  themselves ;  e.  g.,  a 
nitro-protein  made  by  treating  rabbit  serum  protein  with  nitric  acid, 
will,  if  injected  into  even  the  same  rabbit,  cause  the  formation  of 
antibodies  which  will  react  with  this  same  nitro-protein,  and  also 
with  nitro-proteins  derived  from  entirely  different  species  or  even 
from  plants, — but  it  reacts  only  with  nitro-proteins.  It  is  also  possi- 
ble to  cause  chemical  modifications  analogous  to  the  physical  modi- 
fications previously  mentioned,  which  change  only  the  scope  of 
specificity  of  the  antigen  without  altering  its  specificity  for  species. 
A])preciating  that  the  number  of  different  aromatic  radicals  in  the 
protein  molecule  is  not  sufficient  to  account  for  the  innumerable 
manifestations  of  specificity.  Pick  interprets  the  significance  of  these 
aromatic  radicals  as  that  of  a  central  complex  about  which  are  the 
g7-ou])iiigs  wliicli  determine  species  s])ecificity.''-''^  Tt  is  not  merely  the 
number  and  proportion  of  amino-acid  radicals  in  the  jn'otein  molecule 
which  determine  its  specificity,  but,  more  important  because  present- 
ing greater  possibilities  for  variations,  the  arrangement  of  these 
radicals    in    the    molecule.     Contemplating    the    possible    number    of 

infj  substances  imist  liarinoiii/.o,  just  as  tlio  \ilpratioii  of  one  tuniuir  fork  starts 
vibrations  in  anotiier  fork  only  when  Die  two  are  in  liarinonv.  or  as  electroinair- 
netio  waves  incite  resonance  phenomena  (see  Zeit.  f.  Iinmunitiit.,  1911  (0),  246 
and  779). 

■«2a  Landsteiner  and  Prasek  (Zoit.  Innnunitiit..  1913  (20).  211  ),  lic)W(>ver,  state 
that  alteration  of  proteins  by  simply  treatin<f  tliem  with  acid  alcolio!  also  causes 
them  to  lose  their  species  specificity,  and  this  witliout  aiiy  substitution  in  the 
aromatic  radicals  of  the  proteins.  This  observation  throws  doub*  on  the  hypothe- 
sis of  Pick  that  tiic  aromatic  radicals  ai'c  the  essential  center  of  sjx'cics  specificity. 


TOXiyS  AND  ANTirOXIXS  177 

variations  in  the  arrangement  of  the  amino-acids  in  a  protein  which 
the  great  number  of  these  radicals  provides,  there  is  no  difficulty  in 
understanding  the  existence  of  an  almost  limitless  number  of  specific 
distinctions  between  proteins.  Abderhalden,  indeed,  calculates  that 
The  20  amino-acids  we  find  in  proteins  could  form  at  least  2,432, !)02,- 
008,176,640,000  different  compounds,  and  this  without  including  pos- 
sible compounds  varying  in  (luantitative  relations.  A  contribution  to 
the  chemical  basis  of  specificity  has  been  made  by  Kossel,^-''  who  finds 
certain  relations  in  the  proportions  and  groui)ings  of  the  scanty  num- 
ber of  amino-acids  that  make  up  the  protamines  and  histones  of  sperm 
to  be  characteristic  of  the  sperm  of  certain  species  and  families. 

In  the  subsequent  discussion  of  the  various  reactions  of  immunity 
tlie  subject  of  specificity  will  receive  further  consideration.  Of  these 
reactions,  one  of  the  simplest  and  most  studied  is  that  of 

TOXINS  AND  ANTITOXINS 

In  the  preceding  chapter  on  the  bacteria  and  their  products  the 
nature  of  the  true  toxins  was  defined,  and  attention  was  called  to  the 
fact  that  one  of  their  most  important  characteristics  is  that  immuniza- 
tion of  animals  against  them  leads  to  the  accumulation  in  the  blood 
of  substances  capable  of  neutralizing  their  poisonous  action.  Such 
true  toxins  are  produced  especially  by  the  diphtheria  bacillus  and  the 
tetanus  bacillus;  also,  but  less  strikingly,  by  B.  pyocyaneus,  B.  hotu- 
linus,  and  possibly  by  a  few  others.  In  addition  to  these,  numerous 
l)aeteria  produce  hemohjtic  poisons  which  seem  to  have  properties  sim- 
ilar to  the  toxins;  and  there  are  also  toxins  produced  by  plants  (abrin, 
ricin,  crotin,  and  mushroom  poisons)  and  by  animals  (snake  venom, 
scorpion  and  spider  toxin,  and  eel  serum).  Against  all  of  these,  true 
antitoxins  may  be  obtained  by  the  immunization  of  animals. 

Ehrlich's  Conception  of  Toxins  and  Antitoxins. — According  to 
Ehrlicirs  theory,  the  action  of  toxins  upon  cells  is  purely  chemical. 
A  toxin  unites  with  a  cell  because  some  chemical  group  in  the  molecule 
of  toxin  has  a  chemical  affinity  for  some  particular  group  in  the  cell 
protoplasm.  For  convenience  in  description  names  have  been  given 
to  these  groups;  the  group  of  the  toxin  that  combines  with  the  cell 
has  been  called  the  haptopJiorous  group,  or  haptophore,  while  the 
group  in  the  protoplasm  that  combines  with  the  toxin  is  known  as 
the  receptor.*^     It  has  been  found  that  after  being  kept  for  some 

-    42bZeit.  physiol.  Chem.,  1013   (88),  10:3. 

43  Ehiiich  has  used  certain  diatrrams  to  illustrate  these  various  groups  and 
their  relations  to  the  cells  and  to  one  another,  which  are  generally  used  in  ex- 
plaining his  theory.  From  a  teaching  standpoint  they  liave  seemed  to  be  im- 
desirable,  in  that  tlie  student  soon  comes  to  ascribe  physical  properties  and 
appearances  to  wliat  should  I)e  considered  as  chemical  combinations.  The  toxo- 
phore  group  becomes  '"the  black  fringed  end  of  tlie  toxin."  etc.  To  one  accus- 
tomed to  thinking  in  chemical  terms  there  is  no  difficulty  in  following  the  litera- 
ture and  understanding  the  reactions  as  chemical  reactions,  which  thev  arc. 
12 


178  CHEMISTRY    OF    THE    niML'MrY    REACTIOXS 

time,  or  when  placed  under  certain  unfavorable  conditions,  the  toxin 
loses  its  poisonous  properties  without  losing-  its  power  to  combine 
with  cells,  as  shown  by  the  fact  that  immunization  with  such  altered 
toxin  g-ives  rise  to  the  formation  of  antitoxin.  Therefore  it  is  not  the 
haptophore  that  causes  the  harm  to  the  cell,  but  tliere  must  be  some 
other  group  with  this  particular  function.  To  the  group  that  pro- 
duces the  harm  the  name  toxophore  is  given.  If  all  the  receptors  of 
a  cell  are  combined  bj^  toxin  molecules  that  have  lost  their  toxophore 
group  {toxoid  is  the  name  given  to  such  altered  toxins),  the  cell  can- 
not then  be  injured  by  the  corresponding  active  toxin,  showing  that 
the  toxin  must  first  become  united  to  a  cell  receptor  by  its  hapto- 
phore group  before  the  toxophore  group  can  cause  an  injury. 

Animals  that  are  naturally  immune  to  toxins  may  owe  their  im- 
munity to  the  fact  that  their  vital  tissues  contain  no  substances  with 
a  chemical  affinity  for  the  toxin,  and  hence  the  toxin  cannot  unite 
with  them  to  cause  harm.  (In  Ehrlich's  terminology,  the  cells  con- 
tain no  receptors  for  the  toxin.)  The  toxin  may  not  combine  with 
any  tissue  element  at  all  in  such  immune  animals,  and  may  circulate 
for  some  time  harmlessly  in  the  blood,  or  it  may  combine  with  some 
organ  where  it  does  little  harm,  e.  g.,  tetanus  toxin  is  said  to  combine 
chiefly  in  the  liver  of  some  animals,  and  therefore  it  does  not  harm 
their  nervous  system. 

According  to  this  theory,  the  antitoxin  consists  of  cell  receptors 
that  have  been  produced  in  excess  and  secreted  hij  the  cells  into  the 
Mood.  In  the  blood  they  combine  with  any  toxin  that  may  have 
been  introduced,  and  by  saturating  its  affinities  render  it  incapable  of 
uniting  with  the  cells.  As  the  toxin  harms  cells  only  after  it  has 
been  chemically  united  to  them,  it  is  rendered  harmless  when  its 
affinities  for  the  cell  (the  haptophore  groups)  are  saturated  by  cell 
receptors  in  the  blood  stream.  The  process  of  immunization  consists. 
in  injuring  the  body  cells  to  such  a  degree  that  they  are  stimulated 
to  regenerate  the  receptor  groups  with  which  the  toxin  combines; 
these  receptor  groups  are  produced  in  excess,  and  not  only  replace 
those  combined  by  the  toxins,  but  the  excessive  groups  escape  free  into 
the  blood.  Hence  the  serum  of  an  immunized  animal  is  antitoxic 
because  it  contains  free  cell  receptors  that  can  unite  with  the  toxin. 
An  important  point  is  that  the  receptors  liberated  by  all  animals- 
which  have  been  immunized  with  a  given  toxin  seem  to  be  the  same — 
horse  serum,  or  slieej)  serum,  or  goat  serum  will  neutralize  diphtheria 
toxin  if  the  animals  have  been  made  immune  to  this  toxin;  and, 
furthermore,  their  serum  when  introduced  into  the  bod.y  of  an  entirely 
different  animal,  e.  g.,  a  guinea-pig,  will  neutralize  diphtheria  toxin 
within  its  body.  Equally  iinp:)rtaiit  is  the  fact  that  the  antitoxin  for 
one  toxin  will  not  neutralize  any  other  toxin;  e.  g.,  diphtheria  anti- 
toxin will  not  neutralize  tetanus  toxin,  or  conversely.  This  means 
that  diphtheria  toxin  is  attached  to  chemical  groups  of  the  body  cells 


TOXINS  AND  ANTITOXINS  179 

'receptors)  which  are  quite  different  from  the  groups  to  which  tetanus 
toxin  unites,  and  hence  different  receptors  are  thrown  out  in  im- 
munizing against  each.  True  toxins  have  been  designated  monovalent 
antigens,  since  animals  immunized  with  a  purified  toxin  produce  only 
the  one  antibody,  the  antitoxin,  wliereas  many  protein  antigens  pro- 
duce precipitins,  lysins,  agglutinins  and  other  antibodies;  presuma- 
bh^  this  is  because  of  the  relatively  small  size  of  the  toxin  molecule, 
which  limits  the  number  of  its  antigenic  radicals  (Pick).  Or  it  may 
well  be  that  the  immune  body  for  antitoxin  is  quite  different  from  the 
antibod}'  or  antibodies  resulting  from  imnumization  with  non-toxic 
protein  antigens,  for  there  is  much  reason  to  believe  that  the  several 
types  of  reactions  that  may  be  accomplished  with  the  serum  of  animals 
immunized  to  foreign  proteins  or  cells  all  depend  on  one  single  anti- 
body, which  accomplishes  the  destruction  of  the  antigen  by  sensitizing 
it  to  the  enzymes  of  the  blood  and  tissues. 

The  neutralization  of  toxin  by  antitoxin  is  believed  by  many  in- 
vestigators to  be  a  chemical  process,  which  occurs  as  well  in  the  test- 
tube  as  in  the  body.  It  seems  to  occur  according  to  the  laivs  of 
definite  proportion,  a  given  amount  of  antitoxin  neutralizing  a  pro- 
portionate amount  of  toxin  under  equal  conditions  (hence  the  toxin 
is  not  destroyed  by  antitoxin  through  a  ferment  action,  as  was  at 
first  suggested).  Neither  the  toxin  nor  the  antitoxin  is  destroj^ed  in 
the  process  of  neutralization,  as  has  been  proved  by  suitable  experi- 
ments, but  they  appear  to  be  chemically"  united  to  each  other,  as 
any  two  large  molecules  may  be.  Pick  and.Schwarz  believe  that  the 
union  of  toxin  and  antitoxin  takes  place  in  two  steps — first,  colloidal 
adsorption,  and  then  the  specific  reaction.**  There  is  some  question 
as  to  whether  the  union  with  antitoxin  completes  the  neutralization 
of  the  toxin,  or  whether  there  is  then  necessary  a  further  destruction 
of  the  toxin  in  the  body.  But  whether  necessary  or  not,  such  fur- 
ther destruction  does  take  place.  Neutralization  occurs  more  rapidly 
under  the  influence  of  warmth,  and  more  slowly  in  the  cold;  and  it 
is  more  rapid  in  concentrated  than  in  dilute  solutions,  just  as  with 
ordinary  chemical  reactions.  It  is  said  that  it  requires  two  hours 
for  tetanus  toxin  to  be  completely  combined  with  the  corresponding 
quantity  of  antitoxin  at  37°.  According  to  Arrhenius  and  ]\Iadsen, 
reaction  of  antitoxin  upon  toxins  is  accompanied  by  the  liberation  of 
much  heat — 6600  calories  per  gram  molecule,  or  about  half  as  much 
as  is  set  free  by  the  action  of  a  strong  acid  upon  a  strong  base.*^ 
Union  of  toxin  and  antitoxin  causes  no  change  in  the  surface  tension 

4-tAlso  von  Krogh  (Zeit.  f.  Hyg.,  1911  (68),  251).  Bordet.  Biltz..  and  others 
look  upon  the  neutralization  of  toxin   as  an  adsorption  process  entirely. 

4-T  Literature  of  cliemioal  and  physical  reactions  of  toxin  and  antitoxin  triven 
by  Zangger,  Cent.  f.  Bakt.  (ref.),'in05  (36),  2.3S;  Arrhenius,  "Imniuno-ciiem- 
istry,"  1007  and  "Quantitative  Laws  in  Biological  Chemistry,"  London.  Iftl.T; 
also  review  in  Zeit.  Chemother.,  Ref.,  1914  (3),  157;  Oppenheiiner  and  :\Iicliaelis', 
Handbuch  der  Biochemie,  Vol.  II    ( 1 ) . 


180  CHEMISTRY    OF    THE    IMMUXITY    REACTIONS 

of  the  fluid  in  which  the  reaction  occurs  (Zunz),**'  and  the  neutral 
toxin-antitoxin  compound  (diphtheria)  is  not  absorbed  by  animal 
charcoal,  which  absorbs  each  of  the  constituents  when  free.  The 
physico-cliemical  studies  of  the  reaction  between  tetanolysin  and  its 
antibody  gave  results  which  led  Arrhenius  to  conclude  that  in  the 
reaction  there  are  formed  from  one  molecule  of  toxin  and  one  molecule 
of  antitoxin,  two  molecules  of  the  reaction  products  (analogous  to  the 
reaction  between  alcohol  and  acid  which  yields  one  molecule  of  ester 
and  one  of  water).  In  general,  the  union  of  toxin  and  antitoxin  is 
dissociated  by  acids.*'  On  dilution  of  a  neutral  toxin-antitoxin  mix- 
ture, a  certain  amount  of  dissociation  seems  to  occur,  but  there  is  op- 
position to  the  view  that  the  law  of  mass  action  applies  to  the  re- 
action between  toxin  and  antitoxin.  If  toxin  is  added  to  antitoxin  in 
several  fractions,  with  some  interval  of  time  between  each  addition, 
the  final  mixture  is  much  more  toxic  than  if  the  same  quantities  of 
toxin  and  antitoxin  were  put  together  at  one  time.  This  phenomenon 
is  commonlj'  referred  to  as  the  Danysz  effect,  and  indicates  that  the 
toxin-antitoxin  union  is  physical  rather  than  chemical,  for  it  seems  to 
be  quite  analogous  to  such  a  phenomenon  as  the  taking  up  of  more 
dye  by  several  pieces  of  blotting  paper  added  in  series  to  a  dye  solu- 
tion, than  by  the  same  amount  of  paper  added  in  one  piece. 

There  is  no  relation  between  antitoxins  and  enzymes.  The  anti- 
toxin acts  quantitatively,  and  produces  no  detectable  alteration  in  the 
toxin,  or  in  any  other  substance,  as  far  as  we  know.  It  also  has 
but  one  functioning  group  (haptophore),  the  one  with  which  it 
combines  with  the  toxin ;  whereas  both  toxins  and  enzymes  seem  to 
have  two  functionating  groups,  one  which  unites  with  the  cell  or  sub- 
stance that  is  to  be  attacked,  the  other  which  produces  the  chemical 
changes.  But  there  is  evidence  that  union  with  antitoxin  or  fixed  re- 
ceptors prepares  the  toxin  for  its  disintegration,  which,  presumably, 
is  then  accomplished  by  enzymatic  action  as  with  other  antigens. 

CHEMICAL  NATURE  OF  ANTITOXINS 

Tliis  is  as  entirely  unknown  as  is  tlie  nature  of  the  toxins.  In- 
vestigation of  antitoxic  serum  (principally  diphtheria  antitoxin)  has 
shown  that  the  antitoxic  properties  are  closely  related  to  the  serum 
globulin,  which,  however,  by  no  means  proves  that  antitoxin  is  serum 
globulin  or  any  other  sort  of  protein.  According  to  Ehrlich's  theory, 
antitoxin  coiisists  of  free  cell  receptors,  and  these  receptors  are  pre- 
sumaljly  simple  chemical  groups  which  may  be  but  a  part  of  a  larger 
molecule,  or  they  may  be  entire  protein  molecules.  In  any  event 
they  behave  as  colloids;  moving  toward  the  cathode  in  an  electrical 
field,*^  difiPusing  little  or  not  at  all,  their  reaction  curve  resembling 

40  Bull.   Acad.    Roval   ]\red.    Belg.,    1!)II:    also   Bortolini,    BicuOiom.    Zoit..    1010 
(28),  60. 
47Morgenroth  and  As*c1icr.  Cent.  f.  Bakt..  1011    (50),  rAO. 
48  According   to    Field    and    Toaprne    (.lour.    l':\por.    Mi'd..    1007     (0),    80)     both 


CHEMICAL  XATURE  OF  aMITOXIXS  181 

more  an  absoi'ption  curve  than  the  reaction  curves  of  crystalloids,  and 
being  influenced  by  all  conditions  that  influence  colloids.  "Whether 
the  receptor  groups  are  secreted  in  a  free  condition  in  antitoxin  for- 
mation, or  combined  in  a  large  molecule,  is  unknown. 

By  saturating  serum  with  magnesium  sulphate,  or  half  saturation 
with  ammonium  sulphate,  three  chief  groups  of  proteins  can  be  pre- 
cipitatctl  and  isolated.^"  These  are  fibrinogen,  euglobulin  (true 
globulin),  and  pseudo-globulin  (soluble  in  water).  Pick^''  found  that 
the  precipitate  obtained  by  36  per  cent,  volume  saturation  with  am- 
monium sulphate  contained  no  antitoxin ;  the  antitoxin  came  down 
in  the  precipitate  obtained  on  raising  the  strength  from  above  38 
per  cent,  to  46  per  cent.'^^  According  to  Pick,  in  horse  serum  the  anti- 
toxin is  associated  with  the  pseudo-globulin,  and  Gibson  and  Banzhaf 
found  that  the  blood  of  hoi^ses  immunized  to  either  diphtheria  or 
tetanus  toxin  shows  a  marked  increase  (40  to  114  per  cent.)  in  serum 
globulin,  varying  somewhat  according  to  the  antitoxin  content,  the 
more  soluble  globulins  being  most  increased.  At  the  same  time  the 
serum  albumin  and  euglobulin  content  decreases  in  proportion,  while 
the  fibrinogen  shows  no  characteristic  alterations.^-  Hurwitz  and 
Mej^er^^^  however,  find  in  their  study  of  the  blood  proteins  during  im- 
munization, that  the  proportion  of  globulins  increases  according  to  the 
severity  of  the  intoxication,  and  not  in  any  definite  relation  to  the 
degree  of  immunity.  The  average  antitoxic  horse  serum  contains  12 
per  cent,  albumin,  78  per  cent,  of  soluble  globulin  containing  anti- 
toxin, 10  per  cent,  euglobulin.^  By  heating  12  hours  at  57°  a  consid- 
erable part  of  the  soluble  globulin  becomes  insoluble,  without  a  corre- 
sponding loss  of  antitoxin  (Banzhaf). 

The  relation  of  antitoxins  to  proteins  has  also  been  investigated 
by  permitting  digestive  enzymes  to  act  on  antitoxic  serum.  Pick  di- 
gested the  antitoxin-containing  globulin  of  horse  serum  for  several 
days  with  trypsin ;  after  five  days,  when  part  of  the  protein  was 
still  not  digested,  the  antitoxin  was  but  little  impaired  in  strength; 
after  nine  days,  when  most  of  the  protein  was  digested,  the  antitoxin 
had  lost  two-thirds  of  its  strength.  This  indicates  a  considerable  re- 
sistance of  antitoxin  to  tryi3sin,  but  also  shows  that  it  is  affected  in 
much  the  same  way  as  the  globulin   (which  is  itself  very  resistant 

toxin  and  antitoxin  move  towards  the  cathode,  which  is  opposed  to  the  theory 
tliat  tliis  reaction  is  simplv  one  of  oppositely  charged  colloids.  (See  also  Bech- 
hold,  Miinch.  med.  Woch.,  1907    (54),  1921.)" 

49  See  resume  bv  Gibson,  Jour.  Biol.  Chem.,  1905  (1).  IGl  :  GibsoTi  and  Ban/.liaf, 
Jour.  Exper.  Med.,   1910    (12),  411. 

50  Hofmeister's  Beitr.,  1901    (1),  351. 

51  C4ibson  and  Collins  (Jour.  Biol.  Chem.,  1907  (3),  233)  question  the  reliabil- 
ity of  some  of  Pick's  results,  and  repudiate  the  salt  fractionation  metiiod  of  clas- 
sifying proteins. 

52  During  immuniyation  the  antitryptic  power  of  tlie  horse  serum  increases 
with  the  antitoxin  increase  (Krause  and  Klug,  Berl.  kiln.  \\(Hh..  1908  (45), 
1454. 

53  Jour.  Exp.  Med.,  1916    (24),  515:   1917    (25),  231. 


182  CHEMIfiTRY    OF    THE    IMMUNITY    REACTIONS 

to  trypsin)  and  therefore  is  presumably  of  similar  nature.  Anti- 
toxin seemed  to  be  much  more  rapidly  destroyed  by  pepsin-HCl  di- 
gestion than  by  trypsin,  in  which  respect  it  again  resembles  the  serum 
globulin. 

In  favor  of  the  view  that  antitoxin  is  a  definite  protein  body  is 
also  the  fact  that  it  is  not  carried  down  in  indifferent  precipitates,  as 
are  the  enzymes,  but  comes  down  always  in  a  certain  fraction  of  the 
protein  precipitates,  e.  g.,  we  can  precipitate  all  the  serum  albumin 
from  an  antitoxic  senim,  and  it  does  not  carry  down  with  it  any  of 
the  antitoxin.  Another  important  point  has  been  brought  out  by 
Arrhenius  and  Madsen,^*  who  determined  approximately  the  mo- 
lecular weight  of  toxin  and  antitoxin  by  moans  of  their  rate  of  dif- 
fusion, and  found  that  the  toxin  (diphtheria  toxin  and  tetanolysin) 
diffused  ten  or  more  times  as  rapidly  as  the  corresponding  antitoxin. 
Gelatin  filters  also  hold  back  antitoxin  and  let  toxin  pass  through,  and 
toxins  diffuse  into  cells  which  seem  to  be  impermeable  for  the  anti- 
toxin. This  indicates  that  the  antitoxin  molecules  are  much  larger 
than  the  toxin  molecules,  agreeing  with  the  idea  that  antitoxin  is  of 
protein  nature  and  that  toxin  either  is  not  protein  or  is  smaller  than 
most  protein  molecules. 

Taken  altogether,  the  evidence  indicates  a  closer  resemblance  of 
antitoxins  to  proteins  than  has  been  shown  for  the  toxins,  and  all 
attempts  to  separate  antitoxins  from  proteins  have  so  far  failed. 

Antitoxins  are  retained  to  greater  or  less  extent  by  porcelain 
filters,  do  not  pass  through  dialyzing  membrances  readily,  and  are  in 
general  easily  destroyed  by  chemical  and  physical  agencies,  although 
much  less  so  than  are  most  toxins.  Heating  to  60°-70°  injures,  and 
boiling  quickly  destroys  them,  although  like  the  enzymes  and  the  pro- 
teins, they  resist  dry  heat  to  140°,  and  also  extremely  low  tempera- 
ture, without  change.  Putrefaction  of  the  serum  destroys  the  anti- 
toxins (Brieger).^^  They  can  be  preserved  for  a  very  long  time  when 
dried  completely,  but  in  the  serum  they  gradually  disappear,  espe- 
ciall}^  if  exposed  to  light  and  air.  Acids  and  alkalies  destroy  anti- 
toxins, acids  being  the  more  harmful  in  low  concentrations.  Like  the 
enzymes,  antitoxins  are  destroyed  by  ultra-violet  rays.  They  are 
destroyed  in  the  alimentary  tract,  without  appreciable  absorption, 
except  in  the  case  of  new-born  animals  sucking  mothers  whose  blood 
and  milk  contain  antitoxin.^"  Wlien  subcutaneously  injected,  anti- 
toxin  soon  disappears  from  the  blood ;   part  may  be  bound  to  the 

C4  Festskrift,  Siatons  Soniin  Iiislitul.  1902. 

5H  Bf]irin<r  stalos  tliat  totanua  aiiiitoxin  rosists  piitrofnotion. 

66  Rr.mor  and  Much,  .Talirb.  f.  Kindorlioilk..  lOOfJ  (0.3).  fiS4 :  "^rcClintock  and 
Kinp  f.Tf)iir.  Tnfoct.  Dis.,  mOfi  (.3).  701)  found  api)roci:il(lo  absorption  of  antitoxin 
■when  di<,'-cstif)n  was  impaired  by  drufrs.  Full  ivviow  of  literature  on  transmission 
of  antibodies  from  niotlier  to  od'sprins:  sjiven  bv  I'^ainulener,  Jour,  hifeet.  Dis., 
1912  (10),  332:   IbMirlin,  .\reli.  Mens.  Obs!  et  Gyn.,  I!tl2  (1).  407. 


AaaLUTINIXS  A\D  A<;f!IA:ri\.\Tl(>\  183 

tissues,  part  may  be  destroyed,  since  only  traces  appear  in  the  urine. 
It  resists  autolysis. ^^ 

AGGLUTININS  AND  AGGLUTINATION  ''^ 

This  wi'll-kiiown  plienonienon.  the  clumping  or  agglutination  of 
bacteria  when  acted  upon  by  the  serum  of  immunized  or  infected 
animals,  can  hardly  be  considered  as  a  means  of  defense,  since  we 
have  no  evidence  that  it  in  any  way  protects  the  aniraal.^*^  Agglu- 
tinated bacteria  seem  not  to  be  severely  injured  by  the  process, 
and  can  grow  vigorously  in  agglutinative  serum.  Possibly  agglutina- 
tion favors  phagocytosis  and  lessens  dissemination  of  the  infecting 
organisms,  but  it  is  improbable  that  the  influence  on  the  course  of 
infection  is  great.  Agglutination,  therefore,  may  be  looked  upon  as 
an  incident  in  the  infection,  rather  than  as  a  definite  method  of  re- 
sistance, and  it  is  equally  well  produced  hj  immunizing  with  foreign 
cells  or  any  foreign  protein  masses  of  suitable  size  which  contain 
soluble  antigens. 

For  the  production  of  agglutination  it  is  necessary  that  the  cell 
contain  an  antigen  {agglutinogen)  which  has  an  ai^nity  for  the  specific 
constituent  of  the  serum,  agglutinin.  Normal  serum  may  contain 
agglutinin ;  e.  g.,  typhoid  bacilli  are  sometimes  agglutinated  by  normal 
serum,  even  when  it  is  diluted  thirty  times,  but  by  immunization  this 
property  can  be  greatly  increased  until  agglutination  may  be  obtained 
with  dilutions  as  high  as  one  to  a  million.  Whether  normal  agglutinins 
are  essentially  different  from  immune  agglutinins  is  not  known.^® 
Many  protein  solutions,  especially  extracts  of  plant  tissues  and  legum- 
inous seeds,  cause  marked  non-specific  hemagglutination. "''  Likewise, 
bacterial  extracts  may  agglutinate  red  corpuscles.''^  In  immunization 
the  agglutinogen,  which  is  probably  an  intracellular  protein,  acts  as  a 
stimulator  to  the  formation  of  the  specific  agglutinin.  Hence,  when  we 
inject  either  extracts  of  cells  or  entire  cells,  we  secure  agglutinins, 
for  the  agglutinogens  are  liberated  from  the  cells  upon  their  disinte- 
gration.    In  erythrocytes  the  agglutinogen  seems  to  be  in  the  stroma.®^ 

We  can  obtain  agglutinins  against  nearly  all  bacteria,  including 
non-pathogenic  forms,  but  in  varying  strengths.  Agglutinins  are 
found  in  the  blood  stream  in  the  highest  concentrations,  but  they  are 

—    57  Wolff-Eisner  and  Eosenbaum.  Boii.  klin.  Wndi..  lOOfi    (43),  94.5. 

58  Bibliocrraphy  piven  by  Miiller,  Oppenheimer's  Handbuch  der  Bioohemie.  1900 
(II  (1)  ).  .592:  Landsteiner,  ihid.,  p.  428:  Paltanf,  Kolle  and  Wasserniann's  Hand- 
buch., 1913    (II),  483. 

58a  Bull,  however,  would  ascribe  much  importance  to  aff^lutination  of  bacteria 
for  their  removal  from  the  circulation   (.Tour.  Exp.  Med.,  191,5    (22),  484). 

59  See  Andrejew,  Arb.  kaiserl.  Gesundhtsamt.,   1910    (33),  84. 

60  Mendel,  Arch.  Fisiol.,   1909    (7),   168. 

Gi  Fukuhara.  Zeit.  Immunitat.,  1909   (2),  313. 
•■'^  Chyosa,  Arch.  f.  Hyg.,  1910   (72),  191 


184  CHEMLSTRY    OF    THE    IMMUMTY    REACTIONS 

also  present  in  the  various  org-ans,  and  to  greater  or  less  extent  in 
the  other  body  fluids,  excepting  usually  the  spinal  fluid  (Greer  and 
Beeht).®^  The  place  of  their  formation  is  unknown.  Since  bacteria 
contained  within  a  collodion  sac  implanted  in  an  animal  give  rise  to 
the  production  of  agglutinins,  it  is  evident  that  the  agglutinogens 
are  diffusible  to  some  extent,  at  least,  through  collodion.  Old  cul- 
tures of  bacteria  contain  free  agglutinogens,  probably  liberated  from 
disintegrated  cells,  and  filtrates  of  such  cultures  will  neutralize  ag- 
glutinins, showing  both  that  the  agglutinogens  are  filterable,  and 
that  the  reaction  of  agglutination  is  a  eliemical  one  and  not  depend- 
ent upon  the  presence  of  cells.  Agglutinogens  are  said  to  pass 
through  dialyzing  membranes,  while  agglutinins  do  not.  So  it  is  evi- 
dent that  the  agglutinogen  is  of  smaller  molecular  dimensions  than 
tlie  agglutinin,  just  as  toxin  molecules  are  smaller  than  antitoxin 
molecules.  Agglutinogens  are  not  destroyed  by  formalin,  heat,  or 
ultraviolet  rays  in  concentrations  sufficient  to  kill  the  bacteria  con- 
taining them.*'^  Stuber  holds  that  bacterial  agglutinogens  are  lip- 
ine.^*^' 

Properties  of  Agg'lutinins. — Like  most  of  the  other  substances 
produced  in  immunity,  agglutinins  are  precipitated  out  of  the  serum 
in  the  globulin  fraction.  All  attempts  to  separate  them  from  pro- 
teins have  been  unsuccessful.  Stark  ^"  found  that  trypsin  does  not 
attack  the  agglutinins  readily,  corresponding  to  the  resistance  of  the 
serum  globulins  to  this  enzyme ;  alkaline  papayotin  solution  destroys 
them  slowly,  while  pepsin  acts  much  more  rapidly.  Alkalies  are  de- 
structive even  when  quite  dilute,  while  acids  are  much  less  harmful. 
The  temperature  resistance  of  agglutinins  seems  to  be  variable, 
plague  agglutinin  being  destroyed  at  56°,  while  purified  typhoid  ag- 
glutinin may  resist  80°-90° ;  most  agglutinin  serums  lose  their  activ- 
ity at  60°-65°.  The  rate  of  reaction  of  agglutinins  increases  with 
the  temperature,  as  long  as  this  is  not  high  enough  to  injure  the 
reacting  substances."*'  They  are  not  precipitated  by  specific  precip- 
itins, but  are  readily  absorbed  by  charcoal.  They  can  be  formed  by 
spleen  tissue  grown  in  artificial  tissue  cultures."*''^ 

The  structure  of  the  agglutinins  (in  the  Elirlieh  theory)  is  sim- 
ilar to  that  of  the  toxin;  /.  e.,  there  is  a  haptophore  group  by  which 
they  combine  with  the  agglutinogen,  and  a  toxophore  group  by  which 
they  produce  the  changes  that  cause  agglutination.  The  agglutino- 
gen is  probably  related  to  the  antitoxins  in  structure,  having  a  sin- 
gle haptophore  to  unite  w\\h  tlie  agglutinin.  By  degeneration  of  the 
loxo])lior()us  gi'oup  of  tlie   agglutinin,   agglntinolds  nxay  be   formed. 

«3,Toiir.  Infoct.  Dia.,  1910   (7),  127. 

04Stiissano  and  L<'niailc,  Coiii))!.  Kcnd.  Acail.  Sei..  1011    {\'r2) .  (S2?,. 

04a  HioclKMii.  Zoit..,  li)16    (77),  388. 

05  Jnaufr.  Dissert.,  Wiirzhurj,',  lOOo. 

oclMadscn,  ct  nl..  Jonr.  V.\\wv.  Mod.,  1900   (8),  337. 

con  I'ly/gode,  Wien.  klin.  Wocli.,   1913    (20),  841. 


THE  MECHANISM  OF  AOGLUTINATIOy  185 

It  is  believed  that  ag'glutiMins  are  eell  receptors,  wliich  have  a  group 
n'itli  a  chemical  affinity  for  the  agglutinogen  of  the  bacterial  proto- 
plasm, and  also  another  group  which  brings  about  the  agglutination. 
They  are,  therefore,  more  complex  than  the  simple  receptors  that 
unite  with  toxins,  and  are  called  receptors  of  the  second  order.  Ac- 
cording to  Ohno  **'  the  reaction  of  agglutinin  and  antigen  is  in  con- 
stant proportions,  and  seems  to  be  a  chemical  rather  than  a  physical 
reaction.  Coplans  '^^  finds  this  reaction  associated  with  an  increase 
in  conductivity  in  the  solutions,  but  whether  this  depends  upon  the 
agglutinin  reaction  itself,  or  upon  associated  processes,  is  question- 
able. 

Just  what  constituent  of  the  bacteria  acts  as  the  stimulus  to  the 
production  of  the  agglutinin  is  unknown.  Apparently,  there  are 
at  least  two  bacterial  substances  with  this  property,  one  of  which 
seems  not  to  be  a  protein,  since  it  is  soluble  in  alcohol  and  gives  no 
biuret  reaction,  and  resists  temperatures  up  to  165°.  The  other  gives 
all  protein  reactions,  and  is  destroyed  by  heating  to  62°.  We  con- 
sider, therefore,  that  there  are  two  agglutinogens  in  the  bacterial 
cell,  one,  thermostable,  the  other,  thermolabile.  The  difference  in  the 
function  of  these  two  agglutinogens  is  still  a  matter  of  dispute. 
Likewise,  the  question  as  to  whether  they  occur  in  the  membrane  or 
within  the  bacterial  cell  is  still  open,  but  Craw  found  that  the  insolu- 
ble residue  of  crushed  typhoid  bacilli,  after  being  washed  free  of  all 
soluble  constituents,  was  but  slightly  agglutinated  by  active  serum; 
therefore,  the  agglutinogens  are  probably  soluble  intracellular  sub- 
stances. 

Agglutinated  bacteria  can  be  again  separated  from  one  another  by 
the  action  of  organic  and  inorganic  acids,  alkalies,  acid  salts,  and 
by  heating  to  70°  or  75°,  and  after  once  being  separated  they  can- 
not be  reagglutinated  by  fresh  semm.®^ 

The  Mechanism  of  Agglutination. — Tliis  has  been  a  fruitful  field 
of  research,  in  which  the  application  of  physical  chemistry  has  been 
very  profitable.  At  first  it  was  believed  that  the  clumping  was 
brought  about  by  loss  of  motility,  until  it  was  found  that  non-motile 
bacilli  were  equally  affected.  Similarly,  the  hypothesis  of  adhesion  of 
the  flagellfe  was  disposed  of.  Gruber'"  and  others  supposed  that  a 
sticky  substance,  " glahrificm,'"  was  absorbed  from  the  senim  by  the 
bacilli,  which  caused  them  to  adhere  on  contact  with  one  another; 
but  this  does  not  explain  the  flocking  together  of  non-motile  bacilli. 
Paltauf  considered  that  the  specific  precipitin  (see  next  section)  pro- 
duced by  immunization  carried  the  bacilli  down  in  the  precipitate 
formed,  and  there  is  reason  to  believe  that  this  reaction  is  of  im- 

67  Philippine  Jour.  Sci.,  1008   (3).  47. 

fiSJour.  Path,  and  Bact.,  1912    (17).  130. 

eoEisenberg  and  Volk,  Zeit.  f.  Infektionskr.,  1002    (40).  102. 

"0  For  complete  bibliojrraphy,  see  Craw,  Jour,  of  Hygiene.  1005    (.">).  113. 


186  CHEMISTRY    OF    THE    IMMUMTY    REACTIOX^ 

portance,  but  it  does  not  explain  all  the  facts  of  agglutination,  nor  is 
the  relation  between  agglutinating  and  precipitating  power  of  im- 
mune serums  a  constant  one.  In  support  of  this  hypothesis  is  the 
observation  of  Scheller  '  ^  that  mixtures  of  typhoid  bacilli  and  agglu- 
tinating serum  lose  their  agglutinability  by  vigorous  shaking,  which 
may  be  interpreted  as  the  result  of  disintegration  of  the  agglutinating 
precipitate.  Shaking  of  either  'bacteria  or  serum  alone  is  without 
effect.  Neisser  and  Frieduiann  '-  found  that  if  the  bacterial  cells  were 
saturated  with  lead  acetate,  washed  in  water  until  all  soluble  lead  was 
removed,  and  then  treated  with  IToS,  they  were  promptly  agglutin- 
ated and  precipitated,  supporting  other  observations  that  indicate 
that  precipitation  within  the  bacterial  cells  can  lead  to  agglutina- 
tion. This  sort  of  agglutination  is  related  to  the  process  of  formation 
of"  coarse  flocculi  in  solutions,  and  probably  depends  upon  alterations 
in  surface  tension. 

Bordet  and  Gay  described,  under  the  term  conglutination,  the 
observation  that  in  ox  serum  there  is  a  substance  which  combines 
with  corpuscles  (or  bacteria)  that  have  been  acted  upon  by  aggluti- 
nating sera,  and  augments  the  agglutination.'^^  Dean  finds  that,  in 
general,  agglutination  requires  two  agents,  one  being  the  specific 
antibody,  and  the  other  a  precipitable  substance,  probably  a  globu- 
lin. When  cells  have  combined  with  the  antibody  the  precipitable 
substance  is  aggregated  on  their  surfaces  and,  presumably,  determines 
the  agglutination.  Co-agglutination,  described  by  Bordet  and  Gen- 
gou  as  the  agglutination  by  an  antigen  and  the  homologous  antibody, 
of  the  corpuscles  of  another  animal,  is  probably  closely  related  to  these 
phenomena  (Dean). 

Bordet  ^*  made  the  important  observation  that  agglutination  would 
not  occur  if  both  the  bacterial  suspension  and  the  agglutinating 
serum  were  dialyzed  free  from  salts  before  mixing;  but  if,  to  such 
mixtures,  a  small  amount  of  NaCl  was  added,  agglutination  and  pre- 
cipitation of  the  bacteria  occurred  at  once.  This  observation  brought 
the  phenomenon  of  bacterial  agglutination  into  close  relation  with  the 
precipitation  of  colloids  by  electrolytes,  Bordet  comparing  it  to  the 
precipitation  of  particles  of  inorganic  matter  suspended  in  the  fresh 
water  of  rivers  that  occurs  when  the  fresh  water  meets  the  salt  water 
of  the  ocean.  He  found  that  the  agglutinin  combined  with  the  bac- 
teria in  the  absence  of  the  salts,  and  the  resulting  compound  was  pre- 
cii)itated  by  the  addition  of  minute  amounts  of  electrolytes,"'''  which 
alone  did  not  ])recipitate  or  agglutinate  the  bacteria  or  the  serum. 

71  Cent.  f.  Bakt.,  1910   (.'54),   l.'iO. 

"Miinch.  med.  Woeli.,  lOO-l   (51),  4G5  and  827. 

T3  Litoraturo  jrivon  bv  Dean,  Proc.  IJoval  Soc.  (V.) .  1011  (S4),  41(i:  ITall. 
Univ.  Calif.  Publ.,  Pathol.,  1013   (2),  111." 

74  Ann.  d.  I'lnst.  Pasteur.  1800    (1.3),  22.'). 

74a  Corroborated  for  sensitized  red  corpuscles  bv  Eisner  and  Kriedeniann.  Zcit. 
Imniniiiiiit.,   1014    (21),  ,''>20. 


THE  MECII.WISM  OF  ACI(lLrTI\\TfO\  187 

Tlii.s  indicates  that  the  ap'g'hitinins  cause  a  eliaii^e  in  tht-  bacteria  which 
brings  them  under  the  same  physical  laws  as  the  inorganic  colloidal 
suspensions,  which  are  characterized  by  being  precipitated  by  the 
addition  of  traces  of  electrolytes.'^^  This  precipitation  is  imdoubtedly 
due  to  changes  in  solution  tension  and  surface  tension  (see  ''Precipi- 
tation of  Colloids,"  introductory  chapter).  Before  the  agglutinin 
combines  with  the  bacteria  they  behave  like  the  colloidal  solutions 
of  organic  colloids,  lieing  precipitated  only  by  the  salts  of  heavy 
inetals,  alcohol,  formalin,  etc.,  or  by  great  concentrations  of  neutral 
salts.  Field  and  Teague  '^®  have  found  that  agglutinins  carry  positive 
charges  while  bacteria  are  negative,  and  that  by  the  electric  current 
agglutinins  can  be  separated  from  bacteria  with  which  they  have  com- 
bined;  this  shows  that  the  agglutinin  is  not  destroyed  in  the  reaction. 
Teague  and  Buxton  ^^  consider  that  neutralization  of  the  electric 
charge  of  the  bacteria  is  not,  however,  the  only  important  factor  in 
agglutination. 

According  to  Bechhold  ~^  normal  bacteria  behave  like  inorganic 
suspensions  that  have  each  particle  protected  by  an  albumin-like 
membrane,  which  prevents  them  from  being  thrown  out  of  suspen- 
sion by  solutions  of  alkali  salts,  etc.  After  being  acted  on  by  agglu- 
tinin they  are  so  altered  that  they  behave  like  the  unprotected  inor- 
ganic suspensions,  and  are  precipitated  by  salts  and  other  electro- 
lytes. This  suggests  the  possibility  that  the  agglutinin  makes  the 
bacteria  permeable  for  these  electrolytes.  Buxton  and  ShaflPer  "^  also 
found  that  bacteria  which  have  been  acted  upon  by  agglutinin  be- 
have as  if  their  proteins  had  been  so  changed  that  they  are  more 
capable  of  absorbing  or  combining  with  salts  than  when  in  their  nor- 
mal condition.  Strong  salt  solutions  inhibit  agglutination  by  prevent- 
ing the  binding  of  the  agglutinin.^"  Tulloch  ^°^  observed  that  in  the 
presence  of  salts  of  mono-  and  di-valent  cations,  unsensitized  bacteria 
do  not  readily  precipitate  or  agglutinate,  but  sensitized  bacteria,  as 
Bordet  show^ed,  agglutinate  with  small  quantities  of  salts.  In  this 
respect  unsensitized  bacteria  behave  like  "non-rigid  colloids,"  such  as 
fresh  egg  white,  while  sensitized  bacteria  resemble  "rigid  colloids." 
such  as  denatured  egg  white.  Hence  he  advances  the  hypothesis  that 
the  process  of  sensitization  is  akin  to  that  of  denaturation  of  proteins, 
the  specificity  perhaps  depending  on  difiPerent  degrees  of  denatura- 

75  Arrheiiius  (Zeit.  physikal.  Chem.,  in03  (46),  415)  lias  attempted  to  show 
that  the  gas  laws  are  applicable  to  tlie  partition  of  afrslutinin  between  the  bac- 
teria and  the  medium,  which  he  compares  to  tlie  partition  of  iodin  between  water 
and  carbon  disuli)hid.  This  idea  is  not  accepted  by  Craw  (loc.  rit.).  nor  by 
Dreyer  and  Douglas,  Proc.  TJoval  Soc,  1910   (S2),  185. 

76, Jour.  Exper.  INFed.,   1007    (0),  SO. 

77  Zeit.  physikal.  Chem.,  1007    (57).  70. 

78  Zeit.  f.  physikal.  Chem.,  1004    (48),  .385. 

79  Zeit.  physikal.  Chem.,  1007   (57),  47. 

80  Landsteiner  and  St.  Welecki,  Zeit.  Immunitat.,  1910   (8),  397. 
80a  Biochem.  Jour..  1014   (8),  203. 


188  CHEMISTRY    OF    Till;    IMMUyiTY    REACTIONS 

lion.  Agglutination  obeys  the  same  laws  as  other  similar  physical 
phenomena ;  the  rate  of  agglutination  depends  upon  the  concentration 
of  the  suspension  and  of  the  electrolytes,  and  varies  with  the  valence 
of  the  cations.  Although  bacteria  in  an  electric  stream  move  toward 
the  anode  like  all  suspensions,  after  being  acted  on  by  agglutinin  they 
are  agglutinated  by  the  current  between  the  poles ;  ^^  this  indicates 
the  importance  of  the  electrical  charges  of  the  bacterial  surfaces  in 
tlieir  agglutination  reactions. 

In  all  respects  the  behavior  of  bacteria  and  agglutinin  resembles 
the  behavior  of  colloidal  mixtures  in  suspension  (Neisser  and  Friede- 
mann)  ^~  which  form  an  electrically  amphoteric  colloidal  suspension, 
so  that  the  ions  of  electrolytes  or  the  electric  currents,  by  discharging 
them  unequally,  cause  precipitation.  Pliysico-chemical  researches, 
however,  have  yet  failed  to  explain  the  specific  character  of  the  ag- 
glutinins for  specific  bacteria,  but  IMiehaelis  *^  has  developed  an  inter- 
esting analogy  in  the  specific  agglutination  of  bacteria  by  acids.  This 
is  based  on  the  fact  that  the  optimum  concentration  of  H  ions  which 
precipitates  proteins  from  solution  is  characteristic  and  constant  for 
each  protein,  and  the  same  is  true  for  the  agglutination  of  bacteria 
by  acids,  the  agglutination  by  acids  being  even  more  sharply  specific 
in  some  cases  than  the  agglutination  by  immune  sera ;  e.  g.,  typhoid 
and  paratyphoid  bacilli  are  readily  distinguished  because  the  former 
are  agglutinated  by  a  concentration  of  H  ions  from  -4  to  8  X  10"'',  while 
paratyphoids  require  16  to  32  X  10'^,  and  colon  bacilli  are  not  agglu- 
tinated at  all  by  acids.  The  acid  agglutination,  however,  does  not  al- 
ways affect  all  strains  in  the  same  way,  some  strains  which  are  not 
readily  agglutinable  by  antisera  also  resisting  acid  agglutination.®^* 
According  to  Arkwright,®^"  typhoid  bacilli  contain  two  extractable 
proteins  that  are  agglutinated  by  acids,  one  at  3.6  X  10"^  and  the  other 
at  1.1  X  10"'' ;  the  former  seems  to  be  related  to,  if  not  identical  with, 
the  substance  that  is  precipitated  by  immune  serum.  Apparently 
acid  agglutination  of  bacteria  belongs  to  the  same  class  of  reactions  as 
the  coagulation  by  H  ions  of  amphoteric  colloids  of  preponderatingly 
acid  character.  Bacteria  which  have  been  sensitized  by  serum  are 
more  sensitive  to  acid  agglutination  than  are  normal  bacteria.^* 

Alterations  in  the  agglutinability  of  bacteria  are  marked,  e.  g., 
strains  of  typhoid  bacilli  freshly  cultivated  from  human  infections 

SI  Ilcclihold:  liowcver,  Buxton  and  Teag-ue  (Kolloid  Zoitsolir.,  1908,  II,  Suppl. 
2)  state  that  agfjlutinin  bacteria  do  move  towards  tlie  anode,  but  slower  tlian 
normal  bacteria. 

82  Miincli.  med.  \A'ocli.,  1!)04  (51),  M\'->  and  S'27 ;  sec  also  (iiiard-MaiiL:iii  and 
Henri,  C'ompt.  Rend.  Soe.  IJiol.,  ]i)04,  vol.  5fl ;  and  Zanggcr,  Cent.  f.  Hakt!  ( ref. ) , 
ino.5    (.'^(i),  225. 

83  Folia  Serologica,  1911  (7).  1010;  also  Benias.li,  Zeil.  InninuiitiU.,  1!U2  (12), 
268. 

83a  See  Kemper,  .Tour.  Tnf.  Di.s.,  ]!)1G   (18),  200. 

831.  Z,Mt.  Imnuniitiit.,  1014   (22),  .SOO;  Jour.  Ilyg.,  1014   (H).  -'(W. 

84  Krumwiede  and  Pratt,  Zeit.  Immunitiit.,  1013    (10).  7^\'i . 


PRECIPITINS  189 

may  be  practically  inagglutinable  even  by  active  scrum,  but  after  pro- 
longed cultivation  on  media  they  may  or  may  not  develop  agglutina- 
bility.  This  phenomenon  has  not  yet  been  satisfactorily  explained, 
but  it  may  depend  on  an  active  immunity  of  the  bacteria  against  the 
agglutinins.  Such  bacteria  injected  into  rabbits  produce  antisera 
that  will  agglutiiuitc  ordinary  agglutinablc  strains,  but  not  themselves; 
hence  they  do  not  lack  agglutinogens.  They  give  normal  complement 
fixation  reactions,  and  hence  do  not  lack  receptors,  and  they  agglu- 
tinate with  acids  and  chemicals  much  tlie  same  as  ordinary  agglutinable 
strains. ^^'"^ 

PRECIPITINS  *■' 

If  to  a  solution  containing  proteins  we  add  in  proper  proportions 
the  serum  of  an  animal  inmiunized  against  the  same  protein,  a  pre- 
cipitate will  soon  form.  AVliile  not  absolutely  specific,  the  quantitative 
specificity  of  the  precipitin  reaction  is  sufficiently  characteristic  to  be 
of  great  value  in  biological,  bacteriological,  and  medicolegal  work, 
and  it  is  of  importance  to  the  physiological  chemist,  since  it  furnishes 
a  means  of  distinguishing  between  closely  related  forms  of  proteins, 
more  delicate  by  far  than  any  known  chemical  reagent.  The  serum 
reactions  also  prove  that  there  are  sometimes  essential  differences  be- 
tween the  proteins  of  different  species  of  animals,  even  when  by  all 
other  methods  these  proteins  seem  to  be  practically  identical ;  e.  g., 
lactalbumin  of  cow's  milk  is  in  some  respect  different  from  lactal- 
bumin  of  goat's  milk  since  it  produces  a  different  precipitin.  Medi- 
colegally  they  offer  an  accvirate  method  of  determining  the  origin  of 
blood  and  serum  stains,  no  matter  how  old  the  stain  may  be ;  thus 
Hansemann  ***  found  that  material  obtained  from  a  mummy  5000 
years  old  gave  the  precipitin  reaction.-" 

Production  of  Precipitins. — For  the  production  of  the  precipita- 
tion reaction  it  is  necessary  to  have  in  the  substance  used  for  immu- 
nization a  certain  group,  the  precipitinogen,  which  when  injected  gives 
rise  to  production  of  precipitin  by  the  animal.  Apparently  almost 
any  protein  may  act  as  a  precipitinogen  if  injected  into  the  proper  ani- 
mal, but  it  must  he  a  foreign  protein;  rabbit  serum  will  not  produce 
precipitins  if  injected  into  a  rabbit,^^  probably  because  it  is  normally 
present  in  the  blood  of  the  rabbit  and  therefore  does  not  stimulate 
any  reaction;  but  certain  chemical  alterations  in  the  proteins  of  an 
animal,  such  as  heating,  iodizing,  or  partial  digestion,  may  render  them 

84a  Mcintosh  and  McQueen,  Jour.  Hyg.,  1914   (1.3),  409. 

85  For  complete  bibliocraphy  of  the  subject  of  "Precipitins"  see  the  resume  by 
Michaelis,  Oppenheimor's  Handb.  d.  Biochemie.  1009.  II  (1),  ,5.52;  Kraus.  Kolje 
and  Wassermann's  Handb..  1913,  II;  Uhlenhuili  and  Stefl'enhairen.  ihid..  III.  2.i7: 
Zinsser.  "Infection  and  Eesi stance."' 

sGMiinch.  med.  Woch..  1904   (.30),  572. 

87  Not  corroborated  by  Scliniidt.  Zeit.  allor.  Pliysiol.,  1907    (7),  369. 

88  Rarely  a  slight  reaction  against  homologous  proteins  has  been  obtained  {iso- 
precipitins ) . 


190  CHEMISTRY    OF    THE    IMMUNITY    REACTIONS 

SO  different  from  the  normal  proteins  of  the  same  animal  that  they  will 
act  as  an  antigen  when  present  in  the  blood  of  that  animal,  or  an- 
other of  the  same  species,  from  which  they  were  derived.  Of  the 
natural  proteins  of  serum  the  giobulins  are  much  more  active  precipi- 
tinogens than  the  albumins.  In  general  the  more  foreign  the  protein, 
the  greater  the  amount  of  precipitin;  closel}^  related  animals,  e.  g.,  rab- 
bit and  guinea-pig,  produce  little  precipitin  for  one  another's  pro- 
teins. This  indicates  distinctly  that  difference  in  species  depends 
upon  or  is  associated  with  difference  in  chemical  composition  of  the 
proteins.  DiffereJit  sioecies  of  animals  have  very  different  capacity 
for  producing  precipitins,  rabbits  producing  active  sera,  while  guinea- 
pigs  can  produce  but  feebly  precipitating  sera.  Cantacuzene  ^'^  be- 
lieves that  precipitins  are  formed  chiefly  in  the  lymphoid  tissues  and 
bone  marrow,  and  that  the  mononuclear  macrophages  are  most  active 
in  their  formation.*"''  Only  proteins  can  produce  precipitins;  when 
split  to  the  peptone  stage  they  lose  this  property,  but  the  proteins  of 
serum  resist  tryptic  digestion  a  long  time  before  losing  their  precip- 
itinogenic  property,"*^  which  is  destroyed  much  more  quickly  by  pep- 
sin-HCl  mixtures.  The  precipitate  itself  is  very  resistant  to  disin- 
tegrative agencies,  including  putrefaction  (Friedberger),"^  but  is 
soluble  in  dilute  acids  and  alkalies.  It  has  the  power  of  binding 
complement  (Gay^-)  and  if  the  complement  causes  solution  of  the 
precipitate,  poisonous  substances  are  formed  (Friedberger).  Ex- 
cess of  antigen  prevents  the  formation  of  precipitate,  or  redissolves 
it,  but  excess  of  antiserum  has  no  effect.  Since  both  reacting  sub- 
stances are  colloids  tliey  follow  the  laws  governing  other  mutually 
precipitating  colloids,  and  precipitation  occurs  only  when  they  are 
brought  together  in  concentrations  that  lie  within  definite  zones  of 
relative  proportions.  It  is,  of  course,'  perfectly  possible  to  have  a  union 
of  precipitin  and  antigen  without  anj^  visible  precipitate  occurring, 
since  the  product  of  the  reaction  is  not  necessarily  insoluble  under  all 
conditions;  in  this  case  the  occurrence  of  a  reaction  must  be  demon- 
strated by  some  other  method,  e.  g.,  the  complement  fixation  reaction. 
No  precipitins  can  be  secured  against  lipoids  or  other  non-protein  sub- 
stances. Possibly  precipitins  can  be  produced  for  closely  related  sub- 
stances with  molecules  approximating  in  size  the  protein  molecule,  e.  g., 
certain  substances  present  in  supposedly  protein-free  filtrates  of  bac- 
terial cultures.  As  with  the  agglutinin  reaction,  electrolytes  must  be 
present  or  precipitation  will  not  occur.     Neither  the  precipitin  nor  the 

so  Ann.  Inst.  Pasteur.  11)08    (22),  04. 

8»a  Spleen  tissue  eiiltivated  artificially  in  tlie  presence  of  liorse  serum  jiroduces 
specific  precipitins  for  liorse  serum,  and  tissue  from  the  spleon  of  a  guinea  pisr 
that  has  received  injections  of  horse  serum  also  develops  precipitins  for  horse 
serum  wlien  grown  in  cultures   (Przygode,  Wien.  klin.  Wo<h.,  1014   (27),  201). 

90  Fleischmann,  Zeit.  klin.  Med.,  fOGG    (50),  515. 

niCcnt.  f.  Bakt.,  1007    (4.3),  400. 

'••-'See  Univ.  of  falif.  Publ.  Pathol.,  1011    (2),  1. 


f 


PRECIPITINS  191 

antigen  seems  to  be  altered  appreciably  by  the  reaction,  since  when 
either  is  separated  from  tlie  precipitate  it  retains  its  original  prop- 
erties. 

Since  precipitation  of  colloids  is  accompanied  by  or  dependent 
upon  an  aggregation  of  their  particles,  the  precipitin  reaction  is 
closely  related  to  the  agglutination  reaction.  The  amount  of  precip- 
itation obtained  is  much  modified  by  the  amount  of  inorganic  salts 
present,  and,  according  to  Friedemann,'''^  there  is  a  general  resem- 
blance between  the  precipitin  reactions  and  the  precipitations  occur- 
ring when  colloids  precipitate  one  another;  i.  e.,  when  an  amphoteric 
colloid  reacts  with  either  an  acid  or  a  basic  colloid.'**  So  far,  however, 
attempts  to  interpret  the  precipitin  reaction,  as  Arrhenius  has  tried 
to  do,  on  the  basis  of  the  laws  of  physical  chemistry,  have  not  met 
with  much  success  (Michaelis).  We  prefer  the  attitude  of  Krogh,"*^ 
who  states  that  the  colloidal  chemical  part  of  immunological  reactions 
is  to  be  looked  upon  as  only  a  preliminaiy  step  to  the  real  chemical 
process  that  completes  the  reaction  and  gives  it  the  specific  characters. 
As  mentioned  in  the  preceding  section,  agglutination  of  bacteria  is  be- 
lieved to  be  independent  of  the  precipitins,  although  very  probably 
influenced  by  them.  As  with  all  the  other  substances  of  this  class,  the 
precipitins  have  a  haptophore  group  by  which  they  unite  to  the  protein 
molecule,  and  another  group  by  which  they  produce  the  change  re- 
sulting in  precipitation.  When  the  latter  group  is  destroyed  by 
heating  to  72°,  the  precipitin  is  converted  into  a  precipitoid,  which 
possesses  the  property  of  preventing  the  precipitation  of  the  specific 
antigen  by  unheated  precipitin.^*'' 

The  immune  serum  contains  the  precipitin,  which  is  the  passive 
reagent  that  is  throw^n  down  by  a  trace  of  the  immunizing  material 
(precipitinogen).  The  resulting  precipitate  is  the  insoluble  modifica- 
tion of  the  previously  dissolved  precipitin,  and  originates  chiefly  or 
entirely  in  the  proteins  of  the  immune  serum, ''^  according  to  the  work 
of  Welsh  and  Chapman,  especially.  But  as  the  precipitate  is  able  to 
sensitize  anaphylactically,  both  actively  and  passively,  it  would  seem 
that  it  must  contain  both  the  antibody  (which  confers  passive  sensi- 
tization) and  antigen,  to  cause  active  sensitization  (Weil).'''^'^  The 
precipitate  may,  when  of  maximum  amount,  contain  more  nitrogen 
than  corresponds  to  the  entire  euglobulin  of  the  immune  serum,  and 
the  euglobulin  contains  all  the  precipitin,  so  it  seems  probable  that  the 

93  Arch.  f.  Hyg.,  1906   (55),  361. 

94  See  Friedemann  and  Friedenthal,  Zeit.  exp.  Path.  u.  Ther'.,  inOG  (3)  73; 
Iscovesco,  Compt.  Rond.  >Soc.  Biol.,  1906,  Vol.  61,  and  subsequent  volumes. 

94a  .Tour.  Infect.  Dis.,  1916    (19),  452. 

94b  Precipitinogens  are  relatively  resistant  to  moderate  heating,  and  heat«d 
extracts  of  bacteria  are  used  for  precipitin  tests  imder  the  name  thermoprecipi- 
tins.     See  review  by  A.  Ascoli,  Virchow's  Arch.,  1913  (213),  182. 

95  Moll,  Zeit.,  exp.  Path.  u.  Ther.,  1906  (3),  325;  Welsh  and  Chapman,  Proc. 
Royal  Soc.,  B.,  1908   (80),  161;  Zeit.  Immunitat.,  1911   (9),  517. 

95a  Jour.  Immunol.,  1916   (1),  35. 


192  CIIEMLSTIx'V    OF    THE    JMMUMTY    KEACTI0N8 

precipitate  consists  of  more  than  the  precipitin  alone ;  it  ma}''  be  added 
that  the  precipitate  is  always  less  in  amount  than  the  total  giobuliu  of 
the  antiserum."'^  It  is  always  greater  when  the  reaction  is  between 
homologous  antiserum  and  antigen,  than  with  even  closely  related  but 
heterologous  antigens,^'  so  that  the  quantitative  measurement  of  the 
amount  of  i)recipitate  is  of  value  in  applying  this  reaction  to  deter- 
mine the  imture  of  protein  solutions.  The  dilution  of  the  reacting 
solutions  is  of  influence,  however,  for  if  in  too  dilute  solutions  weak 
precipitins  may  fail  to  give  reactions;  with  strong  precipitins  the 
influence  of  dilution  is  nuich  less  (Michaelis). 

According  to  the  source  of  the  protein  used  we  recognize  bacterial 
precipitins,  phyto-precipitins  (for  plant  proteins),'*^  and  zooprecipi- 
tins  (for  animal  proteins).  Although  tissue  extracts,  body  fluids, 
and  exudates  are  generally  used  in  immunizing,  i)urified  constitu- 
ents of  these  protein  mixtures  will  also  excite  precipitin  formation, 
e.  (J.,  we  may  immunize  with  caseinogen  as  well  as  with  milk.  Com- 
plete pepsin  digestion  of  proteins  deprives  them  both  of  their  pre- 
cipitability  and  their  power  to  produce  precipitins,  the  former  prop- 
erty being  lost  first.  Trypsin  seems  to  produce  the  same  effect  more 
slowly.  Heating  to  coagulation — indeed,  heating  in  the  autoclave — 
does  not  destroy  the  precipitinogenous  property  of  proteins,  but  modi- 
fies somewhat  the  reactions  of  the  precipitin  obtained,^  and  precipi- 
tinogen is  destroyed  by  alkalies.  The  specificity  of  precipitinogens  is 
so  modified  by  heating  that  the  precipitins  engendered  by  a  boiled 
antigen  react  with  the  boiled  antigen  and  with  similarly  heated  anti- 
gens from  other  species,  but  not  with  unheated  antigens  even  from  the 
homologous  species. - 

As  proteins  introduced  into  the  stomach  are  normally  destroyed 
before  being  absorbed,  they  do  not  enter  the  blood  and  cause  pre- 
cipitin formation.  However,  as  is  well  known,  eating  of  excessive 
amounts  of  egg-albumen  or  other  easily  absorbed  proteins  may  re- 
sult in  their  passing  the  barriers  and  entering  the  blood  stream,  and 
in  this  way  precipitins  have  been  experimentally  produced.  Pre- 
sumably the  precipitin  reaction  is  a  means  of  throwing  such  foreign 
proteins  out  of  solution  and  rendering  them  harmless.  According 
to  Zinsser  ^  and  others,  the  function  of  the  precipitin  is  to  sensitize 

06  Francescliclli,  Arch.  f.  llyg.,  ]!)07   (00).  207. 

07  Welsh  and  Cliapman,  .Toiir.  Hygieno,  1910    (10),  177. 

98  Literatures  on  precipitins  for  vegetable  proteins  given  bv  Wells  and  Osborne, 
Jour.  Infect.  Dis.,  1!)11    (8),  66. 

1  See  Oberniayer  and  Pick,  who  consider  in  detail  the  efVects  of  various  modilica- 
tions  of  proteins  upon  their  ))o\ver  to  incite  precipitin  formalion  (Wien.  kiln. 
Woch.,  1006  (19),  327).  The  precipitability  of  the  serum,  or  its  power  to  pro- 
duce precipitins,  is  not  affected  by  disease  "(Pribram,  Zeit.  exp.  Path.  u.  Ther.. 
1006    (3).  28). 

-'Schmidt,  Bioclieiu.  Zeit.,  1908  (14),  294;  1910  (24).  4.".;  Zeif.  lininunitiil., 
I!tl2    (i:i),  173;  also  Zinsser,  "Infection  and  Resistance,"  l!»14.  p.  2t;(). 

••'.Jour.  Kxper.  Med.,  1912    (15),  529;   1913    (18),  219. 


,l.\.l/'//17,.l.\7N  O/.'  M.LKItCY  193 

tlie  uufoniHHl  forcijiii  pmrtcins  to  the  difjestive  complement,  a  view 
in  harmony  with  tlie  prevailing  tendeiiey  to  correlate  tlie  inunimity 
reaction  witli  defense  through  enzj'niatic  hydrol^'sis. 

Precii)itiii  appeals  in  the  blood  generally  about  six  days  after  in- 
jection of  the  ])rotein,  but  disappears  after  injection  of  eacli  subse- 
quent dose  of  protein,  to  reappear  again  after  a  somewhat  sliorter 
lapse  of  time.  After  injections  are  stopped,  the  precipitin  disap- 
pears rather  rapidly,  but  never  appears  in  the  urine,  although  it 
may  enter  the  fetal  blood  from  the  blood  of  pregnant  female  animals. 
The  presence  of  precipitins  in  the  blood  does  not  seem  to  prevent 
the  excretion  of  the  foreign  protein  in  the  urine,  nor  are  the  animals 
less  susceptible  to  the  toxic  action  of  the  foreign  protein;  indeed,  the 
reaction  is  even  stronger  in  the  immunized  animals,  and  sometimes 
the  ordinary  dose  becomes  fatal.  Certain  antibodies  are  carried  down 
with  the  precipitates  formed  when  the  serum  containing  them  reacts 
under  proper  conditions  with  an  antiserum ;  e.  g.,  diphtheria  antitoxin 
is  precipitated  when  added  to  the  serum  of  a  rabbit  immunized  to  horse 
serum.  This  is  not  true  of  all  antibodies,  however.^^  As  the  pre- 
cipitates formed  in  the  precipitin  reaction,  when  injected  into  a  guinea- 
pig  make  it  passively  hypersensitive  to  the  protein  used  as  antigen  in 
the  precipitin  reaction,  it  would  seem  that  the  precipitin  and  the 
anaphylactin  are  identical  (Weil),^^  or  at  least  closely  associated. 

Chemical  Properties. — In  its  chemical  nature  precipitin  resembles 
the  "antibodies''  generally,  being  precipitated  in  the  euglobulin 
fraction  of  the  serum/  and  slowly  destroj'ed  by  trypsin,  rapidly  by 
pepsin.  It  cannot  be  separated  from  the  serum  proteins.  The  pre- 
cipitation by  precipitins  is  not  an  enzyme  action,  for  the  precipitins 
are  used  up  in  the  process.  It  apparently  does  not  differ  from  pre- 
cipitations of  colloids  by  other  colloids  of  opposite  electrical  charges, 
except  in  that  the  reaction  is  specific. 

ANAPHYLAXIS  OR  ALLERGY 
In  many  instances  the  injection  of  a  foreign  protein  into  an  ani- 
mal produces  severe,  perhaps  fatal,  intoxication.  With  some  pro- 
teins this  natural  toxicity  is  very  marked, — thus  eel  serum  is  fatal 
to  rabbits  and  dog-s  in  doses  of  0.1  to  0.3  c.e.  per  kilo,  and  foreign 
sera  are  commonly  toxic  to  other  animals;  e.  g.,  fresh  bovine  and 
human  serum  are  quite  toxic  to"  guinea-pigs.  This  so-called  "pri- 
mary" toxicity  is  reduced  or  destroyed  in  most  cases  by  heating  to 
56°  for  30  minutes.*^  Almost  any  non-toxic  soluble  protein,  however, 
may  be  made  toxic  for  animals  by  giving  the  animal  a  small  dose  of 

3a  See  Gay  and  Stone.  Jour.  Immunol.,  1916  (1),  83. 

3b  Jour.  Immunol.,  191 6  (1),  1. 

*Fiuick    (Cent.   f.   Bakt.    ( Ref. ) .   100.5    (36).   744)    states  tliat   if  tlie  precii)itin 
serum  is  very  strono:,  part  of  the  precipitin  comes  down  in  the  pseudoiriobulin. 

4a  The  nature  of  the  toxic  agent  is  unknown,  hut  there  is  reason  to  helieve  that 
it  is  formed,  at  least  in  part,  during-  the  coagulation  of  the  drawn  blood. 
13 


194  CflEMlsTh'Y    OF    THE    JilMiMrV    REACTIOXS 

this  same  protein  at  least  eight  days  previous!}'.  This  preliminary 
dose,  which  may  be  extremely  minute,  renders  the  animal  hypersen- 
sitive to  the  same  protein,  so  that  a  relatively  small  quantity  (a  few 
milligrams  in  the  case  of  the  guinea-pig)  of  an  ordinary  entirely 
harmless  protein,  such  as  egg  white  or  milk,  produces  violent,  often 
fatal,  symptpms  when  introduced  into  the  blood  of  the  animal.  We 
have  not  the  space  to  discuss  the  general  features  of  the  reaction, 
its  history  and  its  relation  to  biology  and  pathology,  which  are  fully 
covered  in  many  easily  accessible  reviews,-'  but  shall  limit  our  consid- 
eration to  the  more  definitely  chemical  aspects  of  the  reaction.^ 

The  Substances  Involved  (Anaphylactogens) . — So  far  as  now 
kno^vn.  these  are  always  proteins,  and  with  the  exception  of  gelatin  ^^ 
and  a  few  others,  practically  any  soluble  protein  will  produce  sen- 
sitization and  intoxication  of  susceptible  animals,  i.  e.,  almost  any 
soluble  protein  may  be  an  anaphylactogen.  As  with  the  other  immun- 
ity reactions,  observations  have  been  made  which  are  interpreted  as 
indicating  that  non-protein  substances  can  produce  this  reaction,  but 
these  interpretations  are  not  generally  accepted.'"'  It  is  possible,  how- 
ever, for  non-protein  substances  to  combine  with  or  alter  the  pro- 
teins of  an  animal  so  that  they  become  as  foreign  proteins  to  that 
animal,  and  thus  cause  sensitization;  in  this  way  can  be  explained 
apparent  anaphylactic  reactions  to  iodin  and  arsenic  compounds, 
and  other  non-protein  substances.  So  far  as  my  own  experiments 
show,  nothing  less  than  an  entire  protein  molecule  will  suffice,"  the 
products  of  protein  cleavage  all  being  inactive.^  Presumably  the 
inefficiency  of  gelatin  as  an  anaphylactogen  depends  upon  its  de- 
ficiency in  aromatic  radicals,  since  these  radicals  have  been  shown 
(Vaughan,  Obermeyer  and  Pick)  to  be  particularly  r important  in 
immunological  reactions.  It  is  not  necessary  for  a  protein  to  con- 
tain all  the  knowni  amino-acids  of  proteins  to  be  active,  however,  for 
certain  vegetable  proteins  (zein,  hordein,  gliadin)  which  lack  one  or 

5  Doerr,  Kolle  and  Wassermann's  Handbuch,  1913,  Vol.  II:  and  Zeit.  f.  Tm- 
miinitat.,  1010:  (2,  ref.),  49;  also  v.  Pirquet,  Aroh.  Int.  Med..  1911  I  7),  2.'>9; 
Friodmann,  Jaliresbcr.  Erpeb.  Immunitiitfrsch.,  1911  (6).  31;  Schittenliehn.  ibid., 
p.  115;  Hcktoen,  Jour.  Amer.  Med.  Assoc,  1912  (fiS),  1081;  Zinsser,  Areh.  Int. 
Med.,  1915  (16),  223.  Concerning  anaphylaxis  in  man  see  Lonpcope.  Ainer.  .lour. 
Med.  Sci.,  1916    (152),  625. 

6  Many  of  the  chemical  features  of  anaphylaxis  I  have  covered  in  the  foUowinj: 
series  of  articles:  .Tour.  Inf.  Dis.,  190S  (5).  449;  1909  (6).  506;  1911  (S),  66; 
1911  (9),  147;  1913  (12),  341;  1914  (14),  364  and  377;  1915  (17),  259;  1916 
(19),    183. 

6a  Wells,  Jour.  Amer.  Med.  Assoc.  1908  (50),  527;  Jour.  Infect.  Dis.,  1908  (5), 
459. 

<5b  Concerning  lipoids  as  antigens  see  Meyer,  Zeit.  Inimunitiit.,  1914    (21),  654. 

7  Zunz  (Zeit.  Inimunitiit.,  1913  (60),  580),  however,  claims  to  get  typical  re- 
actions with  the  higher  proteoses  from  fibrin,  a  tiling  T  have  never  succeeded  in 
doing  with  proteoses  from  egg  albumen. 

sAbdcrhalden  (Zeit.  physiol.  Chem.,  1912  (81),  314)  states  that  he  has  ob- 
tained a  positive  reaction  with  a  synthetic  polypeptid  containing  14  amino-acid 
molecules,  including  only  leucine  and  glycocoll. 


^.Y.l/'//r/,.lA7.S'  OR  ALLERGY  195 

more  of  such  amino-acids  as  glycocoll,  tryptophane,  or  lysine,  pro- 
duce typical  reactions.  Some  compound  proteins  are  efficient  ana- 
phylactoprens  (mucin,*""  casein)  but  with  alpha-nucleoproteins  which 
have  been  thoroughly  purified  I  have  obtained  only  negative  results ;  **• 
as  also  with  histon  and  nucleic  acid,  the  isolated  components  of  nu- 
eleins.  Bacterial  substances,  extracts  of  plant  tissues,  purified  plant 
proteins,  and  proteins  obtained  from  invertebrates  and  cold-blooded 
vertebrates,  have  all  been  found  to  be  anaphylactogens,  if  they  can 
be  introduced  by  any  means  into  the  blood  or  tissues  in  a  soluble 
unaltered  condition. 

If  the  proteins  are  rendered  insoluble  by  coagulation  they  become 
inert,  but  proteins  which  cannot  be  made  insoluble  by  heating  (e.  g., 
casein,  ovomucoid)  withstand  boiling  temperatures.  Trypsin  destroys 
anaphylactogens  in  just  the  same  proportion  as  it  splits  the  protein 
molecules ;  thus,  globulins  resist  trypsin  longer  than  albumins,  both  as 
regards  coagulability  and  anaphylactic  activity.  Acids,  alkalies  and 
other  chemical  agents  may  modify  the  reactivity  of  proteins  in  propor- 
tion to  the  changes  in  solubility  or  constitution  which  they  produce.^ 

The  amounts  of  protein  necessary  to  produce  reactions  in  guinea- 
pigs  are  very  small.  With  crystallized  egg  albumin  sensitivity  has 
been  produced  with  one  twenty-millionth  of  a  gram  (0.000,000,05 
gm.)  and  fatal  reactions  are  obtained  after  sensitization  with  one- 
millionth  of  a  gram.  No  other  animal  seems  to  be  so  sensitive  to  this 
reaction  as  the  guinea-pig,  however,  and  rabbits  and  dogs  require 
larger,  and  in  many  instances,  repeated  doses  to  render  them  ana- 
]5hylactie.  Within  certain  limits  large  doses  are  less  effective  in  sen- 
sitizing guinea-pigs  than  small,  e.  g.,  one  milligram  of  most  proteins 
will  usually  be  much  more  effective  than  one  hundred  milligrams. 
Wliite  and  Avery  ^^  found  that  there  is  a  certain  relation  between  the 
minimum  sensitizing  and  the  minimum  intoxicating  dose ;  with  ex- 
tremely minute  sensitizing  doses  a  larger  intoxicating  dose  is  required 
to  produce  fatal  reaction  than  when  the  sensitizing  dose  is  larger. 

It  is  now  generally  accepted  that  both  the  sensitizing  and  intox- 
icating agent  are  (or  are  derived  from)  one  and  the  same  protein, 
but  the  minimum  intoxicating  dose  is  always  larger  than  the  mini- 
mum sensitizing  dose:  thus,  with  pure  egg  albumin  the  minimum 
lethal  dose  for  sensitized  pigs  was  one-twentieth  to  one-tenth  milli- 
gram by  intravascular  injection,  or  about  one  hundred  times  more 
than  the  minimum  fatal  sensitizing  dose.  With  less  soluble  proteins 
the  disparity  is  even  greater,  for  with  such  the  sensitizing  dose  is  not 

?■!  Elliott,  Jour.  Infect.  Dis.,  1914    (15),  501. 

sb  See  review  in  Zeit.  Immunitat.,  101.3  (10),  500.  conrernino-  alplia-nucleopro- 
teins.  Avhicli  is  the  type  nsiially  desip:natofl  as  "niirlooproteins."  T  have  found 
beta-nnoleoproteins  to  be  more  effective  antigens  (Jour.  Biol.  Cliem.,  1016  (28), 
11). 

9  See  Dold  and  Aoki.  Cent.  f.  Bakt.,  Ref.  Bellage,  1912   (54),  246. 

9a  Jour.  Infect.  Dis.,  1013  (13),  103. 


196  niEMlSTRY    OF    Tin:    niMI  MTV    }{EACTIO\S 

much  changed,  but  the  mininumi  iutoxieatiiig'  close  is  relatively  much 
increased.  Apparently  an  animal  may  be  killed  by  much  less  antigen 
than  is  required  to  saturate  the  antibodies  present  in  its  body  (Weil). 

The  proteins  concerned  must  be  foreign  to  the  circulating  blood  of 
the  injected  animal,  but  they  ma}^  be  tissue  proteins  of  the  same  ani- 
mal (e.  g.,  placenta  elements,  organ  extracts,  lens  proteins)  which 
are  not  normally  present  in  its  blood.  Indeed  it  has  been  claimed 
that  by  injecting  a  guinea-pig  with  the  dissolved  lens  of  one  eye  it 
will  become  sensitized  so  that  it  will  react  to  a  subsequent  injection 
of  the  lens  from  the  other  e^-e.^"  It  is  also  possible  that  vai'ious 
chemicals  may  so  alter  the  blood  proteins  that  they,  too,  behave  as 
foreign  proteins  to  the  same  animal,  rendering  it  sensitive  to  the 
same  altered  proteins  if  they  are  formed  subsequently  by  another 
injection  of  the  chemical  (e.  g.,  iodin,  salvarsan).  In  general,  tis- 
sue proteins  are  less  active  antigens  than  the  proteins  of  the  blood, 
lymph,  and  secretions,  but  even  keratins  may  produce  anaphylaxis 
when  dissolved  ^^  and  positive  results  have  been  obtained  with  pro- 
teins from  mummies. ^- 

The  Poisonous  Agent  ( Anaphylatoxin) . — The  symptomatology  of  the 
intoxication  which  follows  injection  of  the  protein  into  an  animal 
sensitized  with  the  same  protein,  is  such  as  to  leave  no  question  that 
a  poison  is  responsible,  and  this  is  established  as  a  fact  in  several 
ways,  although  as  yet  the  poison  has  not  been  isolated.  As  the  symp- 
tom complex  is  practically  the  same  no  matter  what  sort  of  protein 
is  being  used,  it  would  seem  that  the  poison  must  always  be  the  same 
or  similar — a  striking  and  surprising  fact  in  view  of  the  extremely 
varied  nature  of  the  proteins  capable  of  inciting  anaphylactic  in- 
toxication. Probably  the  poison  is  a  product  of  cleavage  of  the  pro- 
tein by  tissue  or  blood  enzymes,  which  act  only  in  the  presence  of 
the  specific  antibodies  which  unite  the  protein  to  the  enzjnue  (or 
complemejit).  A^aughau  and  his  collaborators  showed  that  i)roteins 
boiled  with  an  alcoholic  NaOlI  solution  might  be  split  into  two  frac- 
tions, one  toxic  and  alcohol-soluble,  the  other  non-toxic  and  insoluble 
in  alcohol.  The  toxic  fraction  gives  all  the  protein  reactions  (except 
that  of  jVlolisch  for  carbohydrates)  and  in  doses  of  8  to  100  mg.  kills 
guinea-pigs  witli  symptoms  practically  identical  with  those  of  ana})hy- 
lactic  intoxication.  The  uniformity  of  the  toxic  effects  with  prepara- 
tions from  different  sorts  of  proteins  suggests  the  existence  in  every 
protein  molecule  of  some  fundamental  toxic  group,  common  to  all  y 
proteins,  the  specificity  residing  in  other  non-toxic  attached  groups. 
This  and  other  observations  led  him  to  the  hypothesis  that  specific 
enzymes  are  develoix'd   in  response  to  the  presence  of  foreign  pro- 

10  IJlilcnlmtli  and  Haendcl,  Zoit.  f.  Tniiminiliit.,  1910   (4),  761. 
JiKriisiiis,    Arch.    f.    Aufrcnlioilk.,    Sujipl.,    1010    (47),    47;    Clougli.    Aib.    kuis. 
GesuiKDitsiuntc,  lt)ll    (;n),  431. 

12  Ulilcnliuth,  Zeit.  f.  Iminuiiitiit.,  1!)10   (4),  774. 


4A'.l/'//17..1A7.s'  on  ALLKIiOY  197 

teius  in  the  blood  stream,  and  that  upon  injection  of  a  second  dose 
of  the  same  protein  these  enzymes  at  once  disintegrate  it,  and 
some  of  the  cleavage  products  being  toxic  the  anaphylactic  in- 
toxication results.  ]\Iany  of  the  later  developments  in  this  field, 
especially  Abderhalden's  studies  on  "protective  ferments,"  have 
added  support  to  this  hypothesis,  so  that  in  its  fundamental  concep- 
tions it  is  now  the  most  generally  accepted  explanation  of  the  processes 
involved  in  anaphylaxis.^^ 

Friedberger  caiTied  the  matter  a  step  farther  by  showing  that  if 
serum  from  a  sensitized  animal  is  incubated  for  a  short  time  with  the 
same  protein,  and  in  the  presence  of  enough  complement,  a  poison 
is  developed  which  produces  the  typical  symptoms  of  anaphylactic 
intoxication  when  injected  into  guinea-pigs.  Tliis  poison  resists  heat- 
ing at  56°,  but  not  at  65°,  and  is  not  a  true  toxin,  for  it  will  not 
produce  an  antitoxin  immunity.  In  the  absence  of  complement,  or 
when  the  complement  fixation  is  prevented  by  strong  salt  solution,^* 
the  poison  (anaphylatoxin)  does  not  develop,  so  that  the  anaphylactic 
reaction  falls  into  the  same  class  as  the  lytic  reactions,  in  which  the 
non-speeiMc  serum  complement  is  united  to  a  cell  by  the  specific  am- 
boceptor, and  then  causes  lysis  of  the  cell ;  in  anaphylaxis  not  an 
organized  cell  but  a  complex  protein  molecule  is  disintegrated  by  the 
complement,  but  in  either  case  a  poisonous  substance  may  be  liberated. 

This  agrees  with  Vaughan's  hypothesis  in  ascribing  the  poisoning 
to  products  of  protein  disintegration  formed  by  enzyme  action,  but 
differs  in  that  specific  intermediary  substances  or  amboceptors  are  sup- 
posed to  be  developed  by  sensitization,  rather  than  specific  enz;ymes. 
Friedberger  is  of  the  opinion  that  many  or  all  the  different  immunity 
reactions  depend  upon  a  single  antibody,  the  different  reactions  merely 
being  different  methods  of  demonstrating  the  presence  of  the  anti- 
body in  tiie  serum.  The  precipitin  reaction  differs  from  the  anaphy- 
lactic reaction,  he  contends,  only  in  that  in  the  latter  the  specific  pre- 
cipitate is  (L'ssolved  by  complement,  yielding  the  anaphylatoxin. 
There  are  many  objections  ^^  to  accepting  this  idea  in  its  entirety, 
which  we  shall  discuss  later,  but  the  formation  of  a  poison  resembling 
that  of  anaphylaxis,  by  a  digestive  action  of  complement  fixed  to  the 
antigen  bj'  the  antibody,  seems  to  be  well  established,  both  as  regards 
in  vitro  and  in  riro  reactions. 

It  would  seem  probable  that  proteins  may  yield  a  similar  poison  in 
whatever  way  their  hydrolysis  is  brought  about,  provided  the  cleav- 
age is  not  too  deep-seated.     For  example,  Rosenow  ^®  has  found  that 

13  See  Vaiighan,  Amer.  Jour.  Med.  Sci.,  1913  (145),  161;  Zeit.  Immunitiit.,  1011 
(9),  458.     Also  a  full  review  in  his  "Protein  Split  Products,"  Philadelpiiia,  191.3. 

14  Fricdherger's  explanation  of  the  inhibiting  efieot  of  salt  as  interference  witli 
complement  action,  has  been  questioned.  (See  Zinsser,  Arch.  Int.  Med  1915 
(16),  238.) 

13  See  Besredka  et  a/.,  Zeit.  Immunitiit.,  1912   (16),  249. 
16  Jour.  Infec.  Dis.,  1912    (11),  94  and  235. 


198  CHEMISTRY    OF    THE    niMiyiTY    REACTIOXS 

pneumoeocci  and  other  bacteria,  permitted  to  autolyze  for  a  proper 
length  of  time,  produce  poisonous  substances  with  all  the  toxicologic 
characters  of  the  anaphylatoxin.  Too  extensive  autolysis  again  de- 
stroys the  poison,  which  is  also  produced  by  digestion  of  pneumo- 
eocci with  serum  from  normal  guinea-pigs,  and  more  rapidly  with 
serum  from  sensitized  animals,  which  likewise  causes  a  demonstrably 
more  rapid  proteolysis.  The  pneumococcus  anaphylatoxic  poison  is 
soluble  in  ether  and  seems  to  be  a  base,  containing  amino-acids,  but 
Friedberger  did  not  find  anaphylatoxin  made  from  serum  proteins  to 
be  soluble  in  ether  or  alcohol,  nor  was  it  precipitated  with  the  globu- 
lins. The  so-called  "Abderhalden  method"  of  sero-diagnosis  of  preg- 
nancy, which  depends  on  the  presence  of  specific  proteolytic  properties 
in  the  blood,  is  an  especially  studied  instance  of  these  principles,  and 
is  discussed  later. 

Presumahly  anaphylactic  intoxication  is  hut  an  exaggeration  of  the 
normal  process  of  defense  of  the  body  against  foreign  proteins  (in- 
cluding bacteria)  through  digestion.  Normally  this  is  accomplished 
in  the  alimentary  tract,  and  complete  disintegration  past  the  toxic 
stage  is  made  certain  by  the  presence  of  erepsin  in  the  intestinal  wall ; 
but  if  intact  foreign  protein  molecules  reach  the  blood  in  any  way, 
this  same  digestive  destruction  is  performed  by  the  enzymes  of  the 
blood  or  tissues.  So  abnormal  is  the  ''parenteral"  introduction  of 
foreign  proteins  that,  once  it  has  happened,  the  protective  mechanism 
is  stimulated  to  the  production  of  large  amounts  of  proteolytic  sub- 
stances, and  on  this  account  if  another  quantity  of  the  same  protein 
is  again  parenterally  introduced  the  breaking  down  of  the  protein  is 
■extremely  rapid.  Certain  of  the  disintegration  products  are  toxic,  but 
with  the  normal  rate  of  disintegration  the  amount  present  at  any  one 
time  is  inadequate  to  cause  poisoning ;  when  the  proteolysis  is  ac- 
celerated, as  in  the  sensitized  animal,  a  poisonous  dose  may  be  pro- 
duced, with  the  resulting  anaphylactic  intoxication.^"  Whether  this 
proteolysis  takes  place  both  in  the  blood  and  tissues  is  not  known.  It 
has  been  found  that  the  specific  proteolytic  power  of  the  blood  is  in- 
creased in  sensitized  animals,  but  on  the  other  liand,  there  is  evidence 
that  without  the  intervention  of  the  liver  (at  least  in  dogs)  anaphy- 
lactic intoxication  cannot  take  place  (INlanwaring  and  others').  During 
the  reaction,  in  any  event,  ])i'()du('ts  of  jirottMii  liydi-olysis  a])pear  in 
the  blood  (Abderhalden).'^ 

Among  possible  cleavage  products  of  proteins  which  may  be  the 
tf)xic  agent  in  anajiliylaxis,  is  /3-imidazolylethylamine  ("histamine"), 
which  is  derivable  from  liistidine,  as  indicated  by  the  structural  for- 
mulas, and  which  produces  effects  resembling  acute  anaphylactic  in- 

17  Heilner  (Zeit,  Biol.,  1012  (58),  .S.3.3)  bolievos  t.liat  tlio  anaphyhu'tio  poisons 
arc  substances  wliich  normally  arc  dostroyod  by  jirotoolysia,  but  tliat  in  the 
sensitized  animals  there  is  a  depressed  catiibolism  \\hic]i  prcvtMits  tlieir  destruc- 
tion. 

18  Zeit.  physiol.  Chem.,  1912   (82),  100. 


ANAPHYLAXIS  OR  ALLERGY 


199 


toxicatioii/'^  Methylguanidine  is  said  to  produce  somewhat  similar 
symptoms,-"  and  other  amines  possibly  may  be  involved.  (See  Chap- 
ter iv,  Ptomains;  Chapter  xix,  Pressor  liases.) 


CTT 

]  I X     X 

HC=C— CH,— C'T[  ( Nil.,)  —coon 
Histiclinc 


fir 

/^ 
HX     X 

HO=(_'— CH,— CH,— NH, 
/3 — imidazolyletliylaniine 
( ITistainiiu' ) . 


The  relation  of  the  normal  toxicity  of  certain  foreign  sera  to  ana- 
phylactic intoxication  has  not  been  determined,  but  there  seem  to  be 
both  definite  similarities  and  differences,-^  which  have  been  discussed 
by  Loewit;  '-"^  chief  of  these  diflferences  is  the  absence  of  the  bronchial 
spasm  with  pulmonary  emphysema  which  is  characteristic  of  anaphy- 
laxis. 

The  anaphylactic  poison  would  seem  to  be  after  the  order  of  the 
alkaloidal  poisons,  at  least  from  the  pharmacological  standpoint,  since 
it  produces  its  effects  quickly,  and  these  effects,  no  matter  how  se- 
vere, are  strictly  transitory,  passing  off  completely  in  a  few  hours, 
which  indicates  that  (like  morphine,  strychnine,  etc.)  they  do  not 
produce  any  deep-seated  structural  alterations  in  the  tissues.  Ac- 
cording to  Schultz  --  the  chief  effects  are  directly  on  the  smooth 
muscles.  Such  anatomical  alterations  as  are  produced,  of  which 
hemorrhages  and  waxy  degeneration  of  the  voluntary  muscles  of 
respiration  -^  are  most  noticeable,  are  ascribable  to  the  effect  on 
respiration,  which  in  the  guinea-pig  often  amounts  to  total  asphyxia- 
tion through  spasm  of  the  musculature  of  the  bronchioles  (Auer  and 
Lewis)  with  profound  permanent  emphysematous  distension  of  the 
lungs.  This  effect  is  peripheral,  and  is  inhibited  by  atropine.-*  Cal- 
cium salts  also  reduce  anaphylactic  reactions.-'  The  poisonous  frac- 
tion obtained  from  proteins  by  Vaughan's  method  resembles  anaphyla- 
toxin,  in  that  it  causes  a  fall  in  blood  pressure  by  paralyzing  the  vaso- 

19  See  Barper,  "The  Simpler  Natural  Bases,"  London,  1914,  p.  30. 
20Heyde.  Cent.  f.  Physiol.,  1911    (2.5),  441;   1912    (26),  401. 

21  It  lias  been  found  that  extracts  of  various  orsjans  are  especially  toxic  to 
animals,  but  that  this  toxicity  may  be  suppressed  by  a  minute  dose,  for  a  few 
minutes  later  larsre  doses  can  be  injected  with  impunity,  alihoui/h  the  l)lood  of 
the  animal  is  highly  toxic  durinjj  the  immune  period,  which  is  of  brief  duration. 
This  condition  is  called  slcpto-phi/la.ris.  (See  Lambert,  Ancel  and  Bouin.  Compt. 
Rend.  Acad.  Sci.,  1911  (154).  21.)  Vauffhan  reports  the  findinjjr  in  normal  tissues 
of  substances  resembling  his  "protein  poisons,"  which  perhaps  come  from  autolysis 
or  tissue  metabolism  and  may  be  related  to  the  "primary  toxicity"  of  organ  ex- 
tracts. 

2iaArch.  exp.  Path.  u.  Pharm.,  191:?    (73).   1. 

22  Bull.  llvfT.  Lab.,  U.  S.  P.  H.  and  ]\f.  TL  Service,  1912    (80),  1. 

23  See  Cent.  f.   Pathol.,   1912    (23),  945. 

24  Jour.  Exp.  Med.,  1910  (12),  151;  Schultz,  .Jour.  Pharm.  and  Fxi).  Tlier.. 
1913   (3),  299. 

25  Kastle,  Healy  and  Buckner,  Jour.  Infec.  Dis.,   1913    (12),   127. 


200  CIIKMISTJ,')-    OF    THE    niMCMTY    h'EACTJOXS 

motor  endings  in  the  blood  vessels  (Edmunds -"''').  It  also  produces 
local  urticaria  when  rubbed  into  the  skin  and  behaves  much  like 
histamine,  with  which,  however,  it  is  not  identical.  One  gram  of 
casein  yields  enough  of  Vaughan's  poison  to  kill  800  guinea-pigs,  and 
the  poison  seems  to  contain  most  of  the  aromatic  radicals  of  the  pro- 
teins. There  is  also  much  other  evidence  of  the  importance  of  the 
aromatic  radicals'  in  anaphylaxis.-^'' 

Other  effects  of  the  anaphylactic  toxin  are  leucopenia,  eosinophilia,-'' 
reduced  coagulability  of  the  blood,  and  a  severe  fall  of  temperature 
unless  the  dose  of  antigen  is  very  small  when  the  temperature  may 
rise.-^  The  antitrypsin  content  of  the  blood  is  not  increased  in  the 
anaphylactic  animal  (Ando  -'^).  Poisonous  substances  similar  to  ana- 
phylatoxin  appear  in  the  urine  during  the  anaphylactic  intoxication 
(PfeifiPer).-®  Anaphylactic  reactions  are  commonly  associated  with 
jnarked  eosinophilia,  both  local  and  general.-"  As  with  other  poisons, 
anaphylatoxin  produces  different  symptoms  in  different  animals.  In. 
dogs  the  chief  effects  are  a  great  fall  in  blood  pressure,^"  loss  of  coagula- 
l)ility  of  the  blood,  hemorrhagic  enteritis,  but  no  bronchial  spasm.  In 
rabbits  the  heart  is  severely  affected,  while  in  guinea-pigs  there  is  a 
remarkable  lack  of  interference  with  the  heart,  so  that  it  beats  long 
after  respiration  ceases.  A  pressor  substance  has  been  found  in  the 
serum  of  intoxicated  guinea-pigs,  which  is  not  present  in  the  artificial 
anaphylatoxin  and  therefore  presumably  is  produced  in  the  body  of 
the  animal.^^  In  man  the  symptoms  are  most  like  those  in  the  guinea- 
pig.  If  the  protein  is  injected  into  the  skin  of  a  sensitized  animal 
there  follows  a  severe  local  reaction, — hyperemia,  edema,  even  necrosis, 
— indicating  that  in  this  specific  proteolysis,  poisons  are  formed  which 
have  a  profound  local  effect,  especially  on  the  blood  vessels.  Repeated 
anaphylactic  intoxication  may  result  in  structural  changes  in  the 
kidneys,  heart  muscle  and  liver  (Longcope  ^^'').  Metabolism  studies 
may  show  an  increased  toxicogenic  destruction  of  protein, ''""  but  the 
increase  in  amino-acids  presumably  resulting  from  proteolysis  in  the 
sensitized  individual,  is  not  large  enough,  if  it  does  occur,  to  be  demon- 
strated by  chemical  methods.^^'^ 

25a  Zeit.  ImmimitJit.,  1013  (17),  105.  Soe  also  Underliill  and  llciulrix.  .Tour. 
Biol.  Chcm.,  1915    (22),  4G5. 

25b  Seo  Baohr  and  Pick,  Arch.  Exp.  Path.,  1913   (74),  73. 

26  Schlecht  and  Sclnvenkor,   Dent.   Arch.  klin.   Med.,   1912    (108),   405. 

27  See  Vauglian,  et  al.,  Zeit.  Immunitiit.,  1911    (9),  458. 
27a  Zeit.  Iiiinuinitiit,  191:5    (18),   1. 

28  Zeit.  f.  Immunitiit.,   1911    (10),  550. 

21' I^iiteraturc  hy  Moscliowitz,  New  York  INIed.  .Tour.,  -Ian.  7,  1911;  SdiU'cht 
and  Scliwenker,  Arch.  exp.  Patli.  u.  Pharm.,  1912    (68).  Ki.i. 

30  Probahlv  from  influence  U])()n  the  nerve  ondinfrs  of  the  vessels  (Pearce  and 
Eisenhrev,  .Tour.  Infec.  Dis.,   1910   (7),  5115). 

•ii  Hir.sclifeld,  Zeit.  Immunitiit.,  1912   (14).  4G(i. 

3iaJour.  Exp.  Med.,  1913  (18),  G78;  1915  (22).  7!t3 :  also  lUniuhlon.  .lour. 
Immunol.,  1910    (1),   105. 

•'ill.  See  Major.  Deut.  Ardi.  klin.   Me.l..  1!I14    (llti),  248. 

•■•■"•See  .\uer  a!id  Van  Slvke,  Jour.  I'lxj).  .Med..  1913  (18),  210;  P.ar^icr  and 
Dale.   I'.iochcni.  .lour.   1914    (8),  G70. 


.1  \  t/'//r/..i  \/N  ni;  M.i.r.iu.y  201 

There  is,  liowcvcr,  imu-li  (lou])t  as  to  the  identity  of  tlie  process  of 
anaphylatoxin  formation  (as  it  occurs  when  antigren,  antibody  and 
complement  are  incubated  in  vitro)  and  the  process  of  anaphylactic 
intoxication.  In  the  first  place,  a  poisono^^s  character,  apparently 
identical  with  this  "anapliylatoxin,''  may  be  griven  to  serum  without 
the  use  of  any  specific  antibody  whatever;  merely  agitating  fresh 
serum  with  any  finely  divided  foreign  material  that  offers  large  total 
surfaces,  such  as  kaolin,  agar,  or  starch,  is  sufficient,  as  also  is  treat- 
ment with  li])oid  solvents,  such  as  chloroform  (Jobling).  In  fact, 
merely  removing  the  fibrin  from  the  plasma  may  make  the  resultant 
serum  highly  toxic,  even  for  the  very  animal  from  which  it  came. 
Furthermore,  if  anaphylactic  shock  were  the  result  of  anaphylatoxin 
formation  in  the  sensitized  animal  through  the  reaction  of  antigen  with 
antibody  and  complement,  the  intoxication  should  occur  if  antibod}'" 
and  antigen  are  injected  simultaneously  into  an  animal ;  but  as  a  mat- 
ter of  fact  the  animal  receiving  antibody  in  passive  sensitization  will 
not  react  unless  the  antigen  is  injected  at  least  three  hours  after 
the  sensitizing  serum  is  injected. ^^'^  This  incubation  period  is  sup- 
posed to  be  required  for  the  anaphylactic  antibody  to  be  fixed  in  the 
cells  where  the  reaction  takes  place  (Otto),  and  perhaps  in  modifica- 
tion of  the  antibody  so  that  it  has  a  greater  afifinity  for  the  antigen 
than  it  has  while  free  in  the  serum  (Weil)  ^^*';  also  in  acquiring  the 
capacity  to  affect  the  cells  after  union  with  the  specific  antigen.  Fi- 
nally, the  isolated  noustriated  muscle  tissue  (uterus)  of  a  sensitized 
animal  gives  specific  reactions  when  brought  in  contact  with  the 
specific  antigen,  no  matter  how  thoroughly  the  animal's  blood  has 
been  removed  from  the  tissues ;  whereas,  the  uterine  muscle  of  an  ani- 
mal injected  with  sensitizing  immune  serum  only  one  hour  before  kill- 
ing does  not  react  when  in  contact  with  specific  antigen.  Weil  dis- 
putes the  toxic  nature  of  anaphylaxis,  even  in  the  intracellular  reac- 
tion, which  he  calls  a  "cellular  discharge." 

Nevertheless,  the  formation  of  anaphylatoxin  is  an  interesting  phe- 
nomenon which  may  well  be  of  importance  in  human  intoxications, 
even  if  it  is  not  the  essential  phenomenon  of  the  anaphylactic  intoxica- 
tion. So  readily  is  blood  serum  made  toxic  in  vitro  that  it  seems  most 
highly  probable  that  a  similar  development  of  toxicity  may  take  place 
in  the  body.  Jobling  ^^^  has  found  that  intoxication  from  anaphyla- 
toxin formation  seems  to  occur  when  kaolin  is  injected  intravenously 
into  animals,  and  hence  it  is  quite  possible  that  the  presence  in  the 
blood  of  abnonnal,  finely  divided  bodies,  such  as  precipitated  proteins, 
cellular  fragments,  even  bacteria,  may  cause  anaphylatoxin  formation 
in  vivo  just  as  they  do  in  vitro. 

The  mechanism  of  anaphylatoxin  formation  is  not  yet  understood^ 

sidSee  Weil,  Jour.  Med.  Ees..  1014   (30),  87:  Jour.  Tnimuiiol.,  lOlf,    (1),  100. 

sieJour.  Med.  Res.,  1015    (32),   107. 

3if  Jobling,  Petersen  and  Eggstein,  Jonr.  Exp.  Med.,  1015    (22),  500. 


202  CHEMISTRY    OF    THE    IMMUXITY    REACTIOXS 

but  there  is  no  lack  of  theories.  The  original  explanation  was  that 
anaphylatoxin  formation  by  specific  antisera  is  the  result  of  digestion 
of  antigen  in  vitro  by  the  action  of  complement  united  to  the  antigen 
by  the  immune  antibody.  For  the  formation  of  anaphylatoxin  by 
inert  finely  divided  particles,  the  explanation  advanced  was  that  the 
highly  developed  surfaces  of  these  particles  either  activated  comple- 
ment, or  united  it  to  the  serum  proteins  so  that  it  digested  them. 
Jobling^^^  has  advanced  the  hypothesis  that  normal  serum  antifer- 
inents,  which  are  believed  by  him  to  be  lii)oidal  in  nature,  are  bound 
by  the  particles  or  by  specific  precipitates,  so  that  the  complement  is 
free  to  attack  the  serum  proteins.  In  any  case,  it  is  now  generally 
agreed  that  the  poisonous  substance  is  derived  chiefly,  if  not  entirely, 
from  the  serum  and  not  from  the  antigen,  even  in  the  case  of  ana- 
phylatoxin formation  by  specific  antigen-antibody-complement  reac- 
tions.^^"^  Furthermore  it  seems  to  be  the  same,  as  far  as  we  can  analyze 
it  by  pharmacological  methods,  no  matter  what  protein  it  is  derived 
from,  or  whether  manufactured  by  immune  or  by  nonspecific  reactions, 
or  by  chemical  means,  such  as  that  of  Vaughan. 

Jobling,  who  holds  to  the  importance  of  anaphylatoxin  formation 
as  the  cause  of  anaphylactic  intoxication,  presents  the  following  con- 
ception of  anaphylaxis :  During  the  course  of  sensitization  there  oc- 
curs the  mobilization  of  a  nonspecific  protease,  which  is  greatly  in- 
creased during  acute  anaphylactic  shock;  at  this  time  there  is  also  a 
decrease  in  antiferment  which  permits  proteolysis  of  the  animal's  own 
proteins.  As  a  result,  there  is  to  be  found  an  increase  in  noncoagula- 
ble  nitrogen  and  amino-acids  of  the  blood,  and  a  decrease  in  serum 
proteases.  "The  acute  intoxication  is  brought  about  by  the  cleavage 
of  serum  proteins  through  the  peptone  stage  by  a  non-specific  protease. 
The  specific  elements  lie  in  the  rapid  mobilization  of  this  ferment  and 
the  colloidal  serum  changes  which  bring  about  the  change  in  anti- 
ferment  titer." 

The  Anaphylactic  Antibody  (Anaphylactin). — That  anaphylaxis,  like 
other  immunity  reactions,  depends  upon  the  presence  of  sjiecific  anti- 
bodies in  the  blood  of  the  sensitized  animal,  is  sho^\^l  by  the  produc- 
tion of  passive  anaphylaxis  in  normal  animals,  by  injecting  into  them 
a  few  cubic  centimeters  of  blood  or  serum  from  a  sensitized  animal. 
Such  animals  become  sensitive  in  a  few  hours  to  the  specific  antigen, 
no  matter  what  species  of  animal  furnishes  the  serum,  showing  that 
various  anaphylactins  can  unite  with  the  same  complement,  altliough 
strongly  specific  as  to  the  antigen.  In  active  sensitization  the  ana- 
phylactin appears  in  the  blood  in  ap])re('iable  (luantities  about  eight 

siKZeit.   Tnimunilii)..    1014    (2:n,  71:   Jour.  Kxp.  Med..   101.1    (22).  401. 

3iii  1'liat  tlip  antigen  nuist  be  dijiostihlo,  liowin-or,  ia  siijri;osto(l  l)y  the  observa- 
tion of  Ten. Broeok  (Jour.  Biol.  Chem.,  1014  (17),  IM'tO)  that  proteins  raeemi/ed 
by  Dakin's  method,  wliicli  cannot  be  dijjested  by  proteolylie  en/yines.  a)'e  unable 
to  cause  anapliylaxis. 


77//;  .1  \1 /'//!/,.  I  C"/7C'  .WTIIiOhV    i  A  \  A  I'll  YLACTIX)  203 

clays  after  the  sensitizing  injection,  increases  to  a  maximum  between 
the  15th.  and  30th  days,  and  then  very  slowly  decreases.  The  reaction 
of  antibody  and  antigen  is  strictly  quantitative,  as  with  all  ambo- 
ceptor reactions.  Tlie  amount  of  antibody  developed  seems  to  be 
limited,  for  after  a  sensitized  animal  is  given  a  sub-lethal  intoxicating 
dose  of  protein  it  may  be  no  longer  sensitive  to  this  protein,  and  this 
refractory  or  anti-anaphylactic  condition  persists  for  three  weeks  or 
more.  It  has  been  demonstrated,  especially  conclusively  by  Weil  and 
Coca,^^  that  this  refractory  condition  is,  as  Friedberger  suggested,  de- 
pendent upon  saturation  or  exhaustion  of  all  the  anaphylactic  anti- 
bodies, and  hence  the  amount  of  these  antibodies  present  free  in  the 
blood  of  a  sensitized  animal  must  be  relatively  small,  for  a  few  milli- 
grams of  the  specific  protein  is  sufficient  to  saturate  them,  e.  g.,  the 
amount  of  antibody  present  in  3  c.c.  of  serum  from  a  guinea-pig  sen- 
sitized with  horse  serum  could  be  neutralized  with  from  0.0005  to 
0.01  c.c.  of  horse  serum. ^^  They  are,  however,  very  persistent,  remain- 
nig  in  guinea-pigs  through  the  entire  life  of  an  animal  sensitized 
when  young.  They  also  pass  from  the  mother  to  the  fetus,  conferring 
a  passive  sensitization  which,  like  passive  sensitization  from  injec- 
tion of  serum  from  a  sensitized  animal,  is  of  relatively  brief  duration, 
in  contrast  to  the  persistence  of  active  sensitization. ^^'^  Anaphylactin, 
like  amboceptor,  resists  heating  at  56°  for  one  hour,  and  is  salted  out 
from  serum  in  the  globulin  fraction.^^'^  ■  Friedberger  contends  that  it 
is  identical  with  the  precipitin,  a  view  yet  under  discussion,'*  but 
strongly  supported  by  Weil 's  observations.^*^ 

Weil  ^■'  has  observed  certain  phenomena  which  led  him  to  conclude 
that  in  anaphylaxis  the  specific  antibody  must  be  largely  fixed  in  the 
cells,  and  that  it  is  in  the  cells  that  the  reaction  occurs;  the  anti- 
bodies present  in  the  blood  of  the  sensitized  animal  are  insufficient  to 
protect  its  cells  from  the  foreign  protein,  hence  the  cellular  intoxica- 
tion. In  support  of  this  idea  is  the  observation  of  Dale  ^*'  that  the 
isolated  smooth  muscle  of  sensitized  guinea-pigs  is  specifically  sensitive 
to  the  foreign  protein.     Weil  states  that  ' '  all  the  evidence  proves  con- 

32Zeit.  Immiinit-it,  1013    (17),  141. 

33  Anderson  and  Frost,  Jour.  Med.  Res.,  1010   (23),  31. 

33a  The  brief  duration  of  passive  sensitization  presumably  dejiends  on  the  forma- 
tion of  antibodies  for  the  foreign  sensitizing  serum,  constitutintr  tlie  condition  of 
"antiscnsitization"  as  contrasted  with  the  refractory  period  wliich  results  from 
the  exliaustion  of  antil)odies  by  antigen.  (See  Weil,  Zeit.  Immunitiit.,  1013  (20), 
199;    1014   (23),  1.) 

3_3b  However,  Scliiff  and  ]Moore  state  that  in  immune  sera  the  allKunin  fraction 
contains  both  the  agent  that  confers  passive  sensitization  and  llic  constituent 
that  causes  the  "primary  toxicity"  of  foreign  sera.  (Zeit.  Immunitiit..  1014  (22), 
CIO.) 

34  See  Zinsser,  Jour.  Exp.  Med..  1012    (15),  529. 
'34a  Jour.  Immunol..  1916   (1),  1. 

35Jour.  Med.  Research,   1913    (27),  407:    1014    (30).  200-364:    1015    (321,   107. 
3G  Jour.  Pharm..  1013   (4),  167. 


204  CHEMISTRY    o/'    THE    fMMCX/TY    REACTIONf? 

elusively  that  anaphylactic  shock  is  induced  by  reaction  between  an- 
chored antibody  and  antigen,  and  tliat  circulating  antibody  plays  abso- 
lutely no  role  in  its  production." 

The  anaphylactin  shows  quite  the  same  characteristics  of  specificity 
as  the  other  immune  antibodies,"'  in  that  proteins  of  closely  related 
species  tend  to  interact,  while  proteins  of  \ery  distinct  biological  or 
chemical  nature  are  easily  distinguished.  Thus,  guinea-pigs  sensitized 
with  ape  serum  will  react  with  human  serum,  but  not  with  serum 
from  dog  or  ox  or  fowl.  However,  in  the  final  analysis,  the  speci- 
ficity depends  upon  the  chemical  composition  of  the  antigenic  protein, 
rather  than  its  biological  origin,  for  I  have  found  it  possible  to  dis- 
tinguish in  the  hen's  egg  five  distinctly  different  antigens,  and  these 
correspond  to  five  proteins  which  have  been  distinguished  by  chemical 
measures.  Together  with  Dr.  T.  B.  Osborne,  working  with  purified 
vegetable  proteins,  I  have  found  evidence  that  a  single  isolated  protein 
(hordein  or  gliadin)  may  contain  more  than  one  antigenic  radical. ^^ 
As  Osborne  ^^  has  said,  "chemically  identical  proteins  apparently  do 
not  occur  in  animals  and  plants  of  different  species,  unless  thej'  are 
biologically  very  closely  related."  Whether  the  chemical  differences 
that  determine  specificity  are  of  quantitative  nature,  which  can  be 
disclosed  by  analytic  means,  or  whether  they  are  sometimes  dependent 
upon  spatial  relationships  of  the  amino-acid  radicals,  as  Pick  sug- 
gests, remains  to  be  determined.  ]\Iy  own  experience  indicates  that 
usually,  at  least,  proteins  distinguishable  by  anaphylactic  reactions 
also  show  readily  distinguishable  chemical  differences. 

THE  ABDERHALDEN  REACTION 

This  reaction  is  based  upon  ,the  hypothesis  that  the  animal  body 
reacts  to  the  presence  of  foreign  proteins  by  providing  specific  means 
of  destroying  them  through  proteolysis,  and  hence  is  fundamentally 
the  same  as  the  anaphylaxis  reaction  as  conceived  by  Vaughan,  Friede- 
mann,  Friedberger  and  others.  It  differs  from  the  other  reactions  of 
this  class  merely  in  that  the  metliods  used  for  determining  the  proteo- 
lysis are  chemical  rather  than  biological.  The  occurrence  of  a  reac- 
tion is  indicated  by  the  production  of  diffusible  products  of  protein 
hydrolysis,  which  may  be  detected  by  any  one  of  several  methods, 
altliough  most  used  is  "ninhydrin"  (triketohydrindene  hydrate) 
wiiich  reacts  with  any  alpha-amino  acid,  the  resulting  condensation 
compound  being  a  blue  or  violet  color,  or  by  observing  the  change  in 
optical  rotation  that  occurs  in  a  solution  of  i)eptone  under  the  hydro- 
lytic  action  of  the  serum. 

It  has  undergone  nnich  llie  same  scries  of  shifting  explanations  as 
the  other  reactions  of  tliis  ehiss.     At  first,  like  the  other  proteolytic 

;"  See  review  in  Jour.  Tnfoet.  Dis..   191 1    (S).  7:?. 
a«.T()ur.  In  fee.  Dis..   1013    (12),  341. 
3'JHarvev   Leetures,    1010   11. 


THE  MiltF.UIIM.DRX   RKM'TIOS  205 

roat'tioiis,  it  was  assumed  that  the  antigen  was  digested;  but,  as  with 
the  preeipitiii  and  anaphylaxis  reactions,  evidence  was  found  by 
iiuuierous  observers  tiiat  not  the  antigen  but  the  proteins  of  the  im- 
mune serum  are  the  chief  or  sole  source  of  the  cleavage  products.  For 
some  reason,  liard  to  explain,  it  has  always  been  referred  to  as  if  it 
were  the  result  of  the  formation  of  specific  enzymes  which  attacked  the 
antigen,  in  spite  of  the  repeated  demonstration  that  sera  giving  posi- 
tive reactions  can  be  inactivated  by  heat  and  reactivated  by  normal 
jserum,^®^  thus  throwing  it  into  the  class  of  amboceptor-complement 
reactions,  with  which  it  agrees  in  principle. 

Having  been  introduced  first  as  a  method  for  diagnosing  pregnancy, 
on  the  principle  that  in  pregnancy  the  chorionic  cells  of  the  placenta 
enter  the  maternal  circulation  and  as  foreign  proteins  cause  the  forma- 
tion of  specific  "defensive  ferments,"  it  was  at  once  taken  up  as  a 
clinical  procedure,  and  as  a  result  an  enormous  literature  on  this 
reaction  was  rapidly  produced.  Much  of  this  represents  highlj-  un- 
critical work,  largely  from  workers  not  trained  or  experienced  in 
immunological  principles,  and  hence  it  is  not  profitable  to  review  it 
in  extenso  here.  Abderhalden's  own  views  are  given  in  full  in  his 
monographs """  and  there  exist  numerous  critical  reviews.^*"^  The 
status  of  the  reaction  at  this  writing  seems  to  be  as  follows : 

Animals,  or  man,  after  having  foreign  proteins  of  any  sort  enter 
the  blood  stream,  may,  and  commonly  do  show  an  altered  condition 
f)f  their  serum,  whereby  when  their  serum  is  incubated  with  the  anti- 
g:en  under  suitable  conditions  very  minute  quantities  of  the  products 
of  protein  cleavage  may  be  set  free,  and  recognized  when  dialyzed 
away  from  the  digesting  mixture ;  or,  a  measurable  change  in  optical 
rotation  of  the  digestion  mixture  occurs.  However,  perfectly  normal 
sera  may  at  times  cause  a  similar  proteoh'sis,  usually  but  not  always 
less  than  with  the  immune  serum. 

The  digestion  seems  to  involve  chiefly  the  serum  proteins  rather 
than  the  antigen,  although  under  certain  conditions  there  may  be 
some  digestion  of  the  antigen.  Bronfenbrenner  holds  that  the  en- 
zymes exhibit  no  selectivity,  digesting  both  the  antigen  and  the  serum 
impartially.^*"* 

Apparently  the  digestion  is  accomplished  by  serum  complement, 
or  at  least  normal  serum  enzymes,  rather  than  by  any  new-formed 
specific  enzyme,  although  enzymes  set  free  from  the  tissues  have  been 
held  responsible  by  some. 

39a  See  Stephan,  Miinch.  med.  Woch..  1914  (61),  SOI:  Ilrtuptmann,  ibid.,  p. 
1167;  Betteneourt  and  !Menezes,  Conipt.  Rend.  Soc.  Biol.,  1!)1G   (77),  162. 

39b  Emil  Abderhalden,  "Scluitzfermente  des  tierisclien  Orpanismus." 

39c  See  Wallis,  Quart.  Jour.  Med.,  1016  (9),  138;  Bronfenbrenner,  Jour.  Lab. 
Clin.  Med.,  1915  (1),  79;  1916  (1),  573.  Hulton,  .Jour.  Biol.  Chcm.,  1916  (25), 
163. 

39d  Supported  by  Smitb  and  Cook,  .Jour.  Infect.  Dis.,  1916  (18),  14.  De  Waele 
states  tbat  it  is  the  serum  globulin  that  is  digested  (Conipt.  Rend.  Soc.  Biol.,  1914 
(76),  627). 


206  CHEMISTRY    OF    THE    nniUXITY    KEACTIOXS 

The  mechanism  of  the  reaction  is  not  understood.  Jobling  and 
Petersen  have  suggested  that  the  antigen-antibody  combination  may 
adsorb  or  bind  the  antipro teases  of  the  serum,  so  that  the  normal  pro- 
tease digests  the  serum  proteins.  Or  it  may  be  that  union  of  antigen 
and  antibody  activates  the  complement,  or  binds  it  to  the  antibody  so 
that  it  digests  either  the  antibody  or  other  proteins  of  the  seinim. 
It  also  is  suggested  that  enzymes  are  set  free  from  the  tissues  injured 
by  the  specific  protein,  or  by  disease,  which  digest  the  foreign  protein 
or  the  cellular  joroteins  that  maj'  have  escaped  from  the  tissues  into  the 
blood  stream. 

The  reaction  possesses  a  certain  specificity,  but  just  the  degree  of 
this  specificity  has  not  been  agreed  upon.  The  claim  of  Abder- 
halden  ^^^  and  his  followers,  that  it  is  by  far  the  most  specific  of  im- 
munity reactions,  whereby  disintegrati'^n  of  small  amounts  of  any 
given  organ  of  an  individual  can  be  determined  by  specific  reactions- 
between  his  serum  and  that  organ,  with  such  refinement  that  even 
cerebral  localization  is  possible,  is  scarcely  credible.  There  are  so 
many  possible  sources  of  error  in  the  original  technic  that  even  with 
great  care  the  charge  of  incorrect  results  fi-om  incorrect  technic  cannot 
be  escaped,  and  therefore,  those  who  do  not  accept  the  doctrine  of  its 
specificity  are  always  on  the  defensive.  Nevertheless,  so  many  careful 
and  experienced  investigators  have  found  the  original  Abderhalden 
reaction  to  give  at  times  absolutely  non-specific  and  hopelessly  para- 
doxical results,  that  its  diagnostic  value  for  either  clinical  or  scientific 
purposes  must  be  considered  at  present  as  unproved,''^^  whatever  the 
final  decision  as  to  its  standing  as  a  specific  reaction  may  be. 

Serum  treated  with  various  inert,  finely  divided  particles,  such  as 
kaolin,  starch,  silicates,  etc.,  may  acquire  the  property  of  giving  posi- 
tive reactions.  This  is  another  point  of  resemblance  to  anaphylatoxin 
foraiation,  and  against  the  specificity  of  the  reaction,  indicating  that 
the  antigen  merely  acts  as  a'  non-specific  adsorbent. 

By  far  the  most  satisfactory  results  have  been  recorded  in  the 
diagnosis  of  pregnancy  by  means  of  placental  antigen.  This  may  be 
explained  by  the  fact  that  the  protease  activity  of  the  serum  seems  to  be 

39e  A  reply  to  numerous  criticisms  is  given  by  Abderhalden,  Fermentforscliung, 
1916  (1),  351;  this  and  other  numbers  of  this  journal  also  consist  lavjroly  of 
articles  on  the  Abderlialden  reaction. 

39tO.  J.  Elsesser  (Jour.  Infect.  Dis..  mifi  (19),  055),  wirking  in  my  labora- 
tory with  the  purified  vegetable  proteins  of  Osborne,  foimd  tlnit  at  the  best  tlie 
specificity  of  the  reaction  was  less  than  that  of  tlie  anaplivlaxis  reactioji,  and 
there  were  many  absolut<']y  non-specific  and  irrational  reactions.  As  tliese  j)ure 
proteins  furnish  a  much  more  appropriate  material  for  studying  specificity  tlian 
the  tissues  or  sera  commonly  used,  it  would  seem  (liat  the  results  thus  obtaiiied 
are  excellent  prof)f  of  tlie  uncertainty  and  unreliability  of  the  reaction.  Careful 
quantitative  studies  of  tlie  setting  free  of  amino-acids  by  serum  incul)ated  with 
placenta,  liy  Van  Slyke  and  his  a^-^sociates,  also  showed  a  c()m))letc  lack  of  spe- 
cific proteolysis  by  pregnancy  scrum  (Arcli.  Int.  ^led.,  1017  (10),  5fl;  Jour. 
Biol.  Chem.,"l915  (23),  377:  see  also  Hulton.  ibid.,  1010   (25),  103). 


OPSOMNki  207 

increased  in  pregnancy,^"*^  and  hence  the  reaction  with  placenta  is  more 
marked  tlian  witli  the  serum  of  non-jiregnant  individuals.  But  simply 
sliaking  nornud  serum  with  kaolin  or  other  foreign  substances  may 
cause  it  to  give  strong  reactions  with  placenta  antigen  (AVallis) . 

OPSONINS  40 

The  correlation  of  phagocytic  and  serum  immunitj'  was  accom- 
plished when  A.  E.  AV right  showed  that,  before  any  considerable 
phagoej'tosis  of  bacteria  can  take  place,  the  bacteria  must  first  be 
acted  upon  by  serum,  which  in  some  way  prepares  them  to  be  in- 
gested by  the  leucocytes.  The  hypothetical  substances  accomplish- 
ing this  sensitization  of  the  bacteria  were  called  opsonins  by  Wright, 
and  the}'  exist  to  a  certain  extent  in  normal  serum,  being  increased 
by  immunization.  Not  only  bacteria,  but  cellular  elements  in  general, 
including  especially  red  corpuscles,  and  even  unorganized  particles 
(such  as  melanin),*^  are  sensitized  for  phagocytosis  by  opsonins. 
Probably  phagocytosis  by  endothelial  *-  and  other  cells  also  requires 
sensitization  of  the  bacteria  by  opsonins.  Although  there  have  been 
many  expressions  of  the  opinion  that  the  opsonins  are  not  distinct 
antibodies,  but  are  identical  with  agglutinins,  bacteriolytic  ambo- 
ceptors, or  other  antibodies,  there  is  much  evidence  to  the  contrary.*^ 
There  are  two  opsonizing  elements  in  serum,  one  thermostable  and  one 
thermolabile,  it  being  the  former  which  is  increased  during  immuniza- 
tion ;  the  thermostable  element  unites  firmly  with  the  object  whieli  is 
to  be  opsonized,  while  the  thermolabile  element  seems  to  remain  free 
in  the  fluid  (Hektoen)." 

It  would  seem  that  opsonization  and  phagocytosis  constitute  but  one 
of  a  series  of  similar  processes  by  which  foreign  proteins  are  removed 
from  the  blood  and  tissues;  i.  e.,  by  lysis  by  extracellular  enzymes 
when  this  is  possible,  as  it  is  vsdth  simple  protein  aggregates  (albu- 
minolysis)  and  with  some  of  the  more  labile  cells  (hemolysis,  bacterio- 
lysis) ;  but  in  the  case  of  more  resistant  structures,  notable  Gram- 
positive  cocci  and  acid-fast  bacilli,  extracellular  lysis  being  unsuc- 
cessful, these  protein  structures  are  taken  within  the  cells  where  a 

30g  See  Sloan,  Amer.  Jour.  Physiol..   1915    (39),  9. 

40  Bibliography  given  bv  Xeufeld,  Kolle  and  \Yassermann's  Handbuch,  191.3  (2), 
440. 

41  Shattock  and  Dudgeon.  Proc.  Eoyal  Soo.    (B),  1908    (SO).  165. 

42  Briscoe,  Jour.  Path,  and  Bact.,  1907  (12),  66.  See  also  ]\Ian\varing  and  Coe, 
who  found  that  the  Kupffer  cells  can  take  up  onlv  opsonized  pneuinococci  (Proc. 
Soc.  Exp.  Biol.,  1916   (13),  171). 

43  See  Hektoen.  .Jour.  Tnfec.  Dis.,  1909    (6).  7S:    1913    (12),   1. 
44Sawtchenko    (Arch.  Sci.  Biol.,   1910   (15),  145:    1911    (16),   161)    holds  that 

there  are  two  steps  in  phagocytosis:  (1)  Fixation  of  the  bacteria  to  the  leuco- 
cyte because  of  modification  of  surface  tension  by  the  fixative  substance  (opsonin 
or  aniboceptor-coniplement  complex);  (2)  Ameboid  motion  of  the  phagocyte; 
an  entirely  independent  phenomenon.  Neither  phase  of  phagocytosis  can  occur 
in  the  absence  of  electrolvtes. 


208  (lIF.MlSTh'Y    OF    Tin:    IMMIMTY    NEACTIOXS 

greater  concentration  oi"  enzymes  may  destroy  them.  Fundamentally 
serum  bacteriolysis  and  phagocytosis  seem  to  he  the  same — in  each  case 
specific  antibody  sensitization  prepares  the  hacterium  for  lysis  by 
enzymes,  either  inside  or  outside  the  cells  that  fur)iish  the  lytic  enzyme. 
As  yet  nothing-  is  known  concerning  the  cliange  brought  about  in 
the  bacteria  by  the  opsonin,  although  it  has  been  established  that  it 
is  the  bacteria  that  are  modified  and  not  the  leucocytes.  The  chemical 
nature  of  the  opsonins  is  likewise  unknown,  except  that  they  may 
combine  with  certain  inorganic  ions  and  are  then  inert  (Hektoen  and 
Ruediger)  ,-*^  since  addition  of  CaCL,  BaCl.,  SrCL  MgCU,  K.SO^, 
NaHCO.j,  sodium  oxalate  and  potassium  ferroc^-anide,  inhibit  the 
opsonic  effect  of  serum.  On  the  contrary,  calcium  salts  stimulate  the 
phagocytic  effect  of  leucocytes,  salts  of  barium  and  strontium  being 
inactive.*®  In  common  with  other  immune  bodies,  opsonins  are 
thrown  down  in  the  soluble  serum  globulins.^"  They  are  very  sensitive 
to  acids  and  alkalies,  being  destroyed  by  a  concentration  of  n/^  and 
their  maximum  effect  is  at  the  neutral  point.**  However,  treatment  of 
either  the  bacteria  or  the  leucocytes  with  very  weak  acids  or  alkalies, 
increases  the  rate  and  amount  of  phagocytosis  (Oker-Blum ).•*'■*  Op- 
sonins may  be  developed  by  immunizing  against  substances  practically 
free  from  protein,  e.  g.,  melanin  granules. ""^  Injection  of  nuclein 
preparations  may  increase  the  amount  of  opsonin  present  in  the 
blood. °°^  Cholesterol  in  excess  diminishes  phagocytosis,  but  appar- 
ently through  its  action  on  the  leucocytes.^'"'  Both  the  sensitization 
of  bacteria  and  their  ingestion  by  leucocytes,  either  with  or  without 
sensitization,  take  place  in  accordance  with  the  laws  regulating  an 
adsorption  process  (Ledingham,"'"''  Schiitze*"). 

THE  MEIOSTAGMIN  REACTION 

Reaction  of  antigens  with  their  specific  antibodies  results  in  lower- 
ing the  surface  tension  of  the  solution  in  which  the  reaction  occurs, 
which  may  be  demonstrated  by  counting  the  number  of  drops  of  the 
fluid  per  minute,  under  constant  conditions.  Ascoli  and  Izar  ^^ 
worked  out  methods  for  practical  application  of  this  phenomenon, 
giving  it  the  name  of  "meiostagmin  reaction,"  from  the  Greek,  mean- 
ing ''small  drop."  The  numl)er  of  drops  from  a  stalagmometer  is 
counted,  and  an  increase  of  two  or  more  per  minute  is  considered  a 

45  .Tour.  Infect.  Dis.,  190.'')   (2),  120. 

4"  llamburfjer,  Biochem.  Zeit.,  1910   (24),  470;   1910   (2()K  M. 

47  See  Simon  et  al.,  Jour.  Exp.  ]\Ied.,  1906  (8),  G51;  I  li'iiiciiiaiin  ami  (lato- 
wood,  .lour.  Infcc-.  Dis.,   1912    (10),  410. 

48  No-ruclii,  .lour.   Kxp.  Mod.,   1907    (9).  4.)4. 

49  Zeit.   linnnuiitiit.,   1912    (14),  48.5;    Sdiiitzo.  Jour.  llv>:.,   1!>14    (14).  201. 
■'-n  Lcdiii^haiii.   Zeit.   liinnunitiit.,   1909    (.3).   ll'.i. 

■"'Oa  lU'dson,  .lour.   Patli.  and  I5act.,  1914    (19).    I'.H. 

cob  Dewev  and  Nu/.um,  .Tour.   Infect.  Dis.,   1914    (hi).  472. 

•"•oo.Tour.'lIyff.,  1912    (12).  320. 

51  :\Iiincli.  nied.  Woch.,  1910   (57),  (12.  182  and  403. 


THE  KI'irilAMS    h'KACTlOX  209 

positive  reaction,  after  two  hours'  incubation  of  the  reacting  mixture; 
the  increase  is  seldom  above  eight  drops.  This  reaction  is  said  to  be 
sharply  specific  and  extremely  delicate,  detecting  antigens  diluted  \\\) 
to  1  in  100,000,000  or  more.  The  antigens  used  are  soluble  in  alcohol, 
but  their  nature  is  unknown ;  the  antibody  involved  in  the  reaction  is 
referred  to  as  the  meiostagmin,  but  its  relation  to  other  antibodies  is 
likewise  unknown. 

THE  EPIPHANIN  REACTION 

Besides  reduction  in  surface  tension,  other  physico-chemical  changes 
result  from  antigen-antibody  reactions,  including  the  rate  of  diffusion, 
the  osmotic  pressure,  and,  in  consequence,  according  to  Weichardt, 
the  neutral  point  to  phenol-phthalein  of  a  mixture  of  barium  hydrox- 
ide and  sulphuric  acid,  is  also  changed  towards  the  acid  side  by  anti- 
gen-antibody reactions  taking  place  in  the  mixture.'^-  This  phenom- 
enon has  been  utilized  by  Weichardt,  under  the  name  of  "epiphanin 
reaction,"  to  determine  the  occurrence  of  such  interaction  of  antigen 
and  antibody.  The  reaction  probably  depends  upon  absorption  phe- 
nomena, but  the  exact  nature  of  the  change  is  not  yet  understood. 
According  to  Rosenthal,"  the  epiphanin  reaction  is  especially  suitable 
for  demonstrating  cancer  antibodies  and  antigens,  but  Burmeister  ^* 
and  others  have  not  been  successful  with  this  procedure. 

52  See  Weichardt.  Berl.  klin.  Woch.,  1911  (48),  1935;  Rosenthal,  Zeit.  Im- 
nuinitat.,  1912  (13),  383;  Angerer  and  Stotter,  Miinch.  med.  Woch.,  1912  (59), 
2035. 

53  Zeit.  Chemotherapie,  1912   (1),  156. 

54  Jour.  Infec.  Dis.,  1913   (12),  459. 


14 


CHAPTER    VIII 

CHEMISTRY  OF  THE  IMMUNITY  REACTIONS  (Con- 
tinued)—BACTERIOLYSIS,  HEMOLYSIS,  COMPLE- 
MENT FIXATION,  AND  SERUM  CYTOTOXINS 

SERUM  BACTERIOLYSIS  ' 

The  bactericidal  property  of  serum  may  be  shown  by  its  destruc- 
tion of  the  life  manifestations  of  bacteria  without  marked  alteration 
in  their  structure,  or  it  may  be  accompanied  by  dissolution  of  the 
bacterial  cell  {'bacteriolysis).  How  much  of  the  bacteriolytic  process 
is  performed  by  the  serum  itself,  or  how  much  by  the  autolytic 
enzymes  of  the  bacterial  cell,  is  unknown,  but  the  latter  is  probably 
a  factor.  The  bactericidal  property  of  immune  seriTm  has  been  sho^^^l 
to  be  quite  independent  of  the  antitoxic  properties  and  also  to  have 
quite  a  different  mechanism.  This  last  is  shown  in  the  following- 
manner  : 

If  w^e  heat  bactericidal  serum  made  by  immunizing  an  animal 
against  bacteria,  say  the  cholera  vibrio,  at  55°  for  fifteen  minutes,  it 
will  be  found  to  have  lost  its  power  of  destroying  these  organisms. 
Normal  serum  of  non-immunized  animals  is  equally  without  effect 
upon  the  vibrios.  If  however,  we  add  to  the  inactivated  heated  serum 
an  equal  quantity  of  inactive  normal  serum,  the  mixture  will  be 
found  to  be  as  actively  bactericidal  as  the  original  unheated  immune 
serum.  This  phenomenon  is  interpreted  to  mean  that,  by  immuniza- 
tion, some  new  substance  has  been  developed  which,  although  by  itself 
incapable  of  destroying  bacteria,  is  able,  when  united  with  some  sub- 
stance present  in  normal  serum,  to  destroy  bacteria  readily.  The 
substance  present  in  normal  serum  is  also  incapable  of  affecting  bac- 
teria by  itself,  but  needs  the  presence  of  the  substance  developed  by 
immunizing  to  render  it  bactericidal.  Hence  the  hactericidal  prop- 
erty in  this  case  depends  on  two  suhstancrs  acting  together:  one,  de- 
veloped during  immunization  and  therefore  caHed  the  inintioie  hody, 
is  specific  for  the  variety  of  bacteria  used  in  immunization,  and  is  not 
destroyed  by  heating  at  55°.  The  other,  present  in  normal  serum,  is 
not  increased  during  immunization,  is  not  (altogether)  specific  in 
character,  and  is  destroyed  by  heating  at  55°  ;  as  its  action  is  eom- 
plementar}'  to  that  of  tlie  specific  innnune  body,  it  is  called  the  com- 
plement.^ 

1  Review  and  liiljlio^rrapliv  l)v  ^Vfiiller,  Oppeiilieimer's  TTaiull).  d.  Bioelicm.,  1009 
(II    (1)   ),  629. 
-  'J'lie  i)olyiiuc]ear  Icucocyies  also  coniain  l)actoriolytic  agents,  "endolysins,"  of 

210 


AMBOCEPTOR  A^D  COMPLEMENT  211 

It  is  believed  tliat  the  action  of  these  substances  is  as  follows :  The 
immune  body  is,  like  antitoxin,  a  cell  receptor  which  unites  the  bac- 
teria to  the  cell.  It  differs  from  the  antitoxin,  however,  in  that  it 
has  two  affinities,  one  for  the  complement  and  the  other  for  the  bac- 
terial substance.  On  account  of  the  existence  of  the  two  affinities  it 
is  called  an  amboceptor.  Some  serums  contain  such  amboceptors  for 
certain  bacteria  without  previous  immunization,  hence  the  term  im- 
mune amboceptor  is  reserved  for  amboceptors  developed  by  immuniza- 
tion. 

Amboceptor  and  Complement. — The  function  of  the  amboceptor 
is  to  unite  the  bacterial  protoplasm,  to  which  it  is  attached  by  one 
affinity,  to  the  complement  which  it  holds  by  its  other  affinity,  or,  to 
put  it  in  a  more  strictly  chemical  way,  the  addition  of  the  ambocep- 
tors to  the  bacteria  gives  them  a  chemical  affinity  for  complement. 
It  is,  therefore,  an  intermediary  body,  uniting  the  complement  to  the 
bacterial  protoplasm.  The  complement  ^  is  the  substance  that  actually 
destroys  the  bacteria,  in  which  respect,  as  well  as  in  its  susceptibility 
to  heat,  it  resembles  the  enzymes.  Complement  is  present  in  normal 
serums,  and,  as  it  is  not  increased  in  amount  during  immunization, 
it  may  not  be  sufficient  to  satisfy  all  the  amboceptors,  hence  it  may 
be  impossible  to  secure  marked  bactericidal  effects  even  when  many 
amboceptors  have  been  formed.  If  the  complement  in  an  immune 
serum  has  been  destroyed  by  heating,  it  may  be  replaced  by  adding 
normal  serum  from  another  animal,  even  of  some  other  species;  indi- 
cating either  that  the  complement  is  not  absolutely  specific  in  its 
nature,  or  that  quite  the  same  complement  may  be  present  in  the 
blood  of  many  different  animals.  The  origin  of  the  complement  is 
unknown,  but  it  has  been  urged  that  the  leucocytes  are  an  important 
source  of  this  substance,  if  not  its  chief  one ;  ^^  there  is  evidence,  how- 
ever, that  various  organs  and  cells  may  also  produce  complement.^*^ 
Its  most  important  characteristics  are  its  extreme  susceptibility  to 
heat,  and  the  resemblance  of  its  action  to  the  action  of  enzymes.'* 
Hektoen  ^  found  that  it  could  be  made  to  unite  vsdth  Mg,  Ca,  Ba,  Sr, 
and  SO^  ions,  which  rendered  the  complement  (for  typhoid  bacilli  and 
red  corpuscles)  inactive.  ]\Ianwaring  '^  found  that  these  ions  could  be 
separated  again  from  the  complement  by  simple  chemical  precipita- 

a  similar   complex   structure,   but   quite   distinct   from   the   serum   bacti.Tiolvsins. 
(See  Kling,  Zeit.  Immunitat.,  1910   (7),  1). 

3  Review  and  bibliography  by  Noguchi,  Biochem.  Zeit.,  1907    (6),  327. 

3a  Cholera  antiserum  will  produce  the  Pfeiffer  phenomenon  of  lysis  of  cholera 
vibrios  in  animals  made  leucocyte-free  with  thorium,  showing  that  the  presence 
of  the  leucocytes  themselves  is  not  essential.  (Lippmann,  Zeit.  Immunitiit., 
1915   (24),  107.) 

3b  See  Dick,  Jour.  Infect.  Dis.,  1913  (12),  111;  and  Lippmann  and  Plesch, 
Zeit.  Immunitiit..  1913   (17),  54S. 

4  See  Walker,  Jour,  of  Phvsiol..  1906   (33),  p.  xxi. 

5  Trans.  Chicago  Path.  Soe.,  1903    (5),  303. 

6  Jour.  Infectious  Diseases,  1904   (1),  112. 


212  CHEMIfiTRY    OF    THE    IMMUNITY    REACTIONS 

tion.  Acids  stronger  than  COo  and  of  the  higher  saturated  or  un- 
saturated fatty  acid  series,  inactivate  complement  in  strengths  greater 
than  n/^f,,  and  alkalies  are  equally  inhibitiveJ  Ultraviolet  rays 
destroy  complement.''^  Sherwood  "''  has  made  a  study  of  various  sub- 
stances that  maj"  be  present  in  the  blood  in  excessive  amounts  during 
pathological  conditions,  such  as  CO,,  lactic  acid,  acetone,  etc.,  and  finds 
that  they  interfere  seriously  with  the  action  of  complement,  wliich 
suggests  that  they  may  favor  infection  or  interfere  with  recovery 
from  infection. 

Presumably  the  complement  is  a  protein,  for  it  has  antigenic  prop- 
erties, so  that  immunization  with  sera  containing  either  complement 
or  complementoid  causes  anticomplement  activity  in  the  blood  of  the 
immune  animal.  Also,  it  is  destroyed  b^^  tiypsin  free  from  lipase,^ 
and,  like  other  colloids,  is  readily  adsorbed  by  surfaces;  like  enzymes, 
complement  is  destroyed  by  shaking,''  and  gradually  disappears  on 
standing.  There  are  some  striking  resemblances  between  the  be- 
havior of  complement  and  of  certain  compounds  of  protein  with  soaps 
and  lipoids,  as  pointed  out  especially  by  Noguehi,  but  that  these  are 
identical  with  true  complement  is  doubtful.  (See  Hemolysis.)  Its 
colloid  nature  is  attested  by  the  large  loss  when  complement  is  filtered 
through  Berkefeld  filters.^"  A  careful  review  of  the  evidence  has  led 
Liefmami  "  to  the  conclusion  that  the  reaction  of  complement  to  sen- 
sitized corpuscles  is  more  like  that  of  ferment  to  substrate  than  of 
antigen  to  antibody. 

According  to  the  Ehrlich  theory,  complement,  like  toxins  and  en- 
zymes, possesses  at  least  two  groups :  one,  the  haptophore,  with  which 
it  unites  with  the  amboceptor;  the  other,  the  toxophore  (or  zymo- 
phore, because  of  its  enzyme-like  action),  which  attacks  the  bacterial 
protoplasm.  It  may  degenerate  and  lose  its  toxophore  group  while 
retaining  the  power  to  combine  by  means  of  its  haptophore  group, 
thus  forming  a  complementoid.  Complement  and  amboceptor  exist 
side  by  side  in  the  serum,  not  uniting  with  one  another  until  the 
amboceptor  has  become  attached  to  the  bacterial  protoplasm. 

It  is  generally  stated  that  if  serum  containing  complement  be  so 
treated  as  to  separate  the  globulins  from  the  albumin,  it  is  found 
that  the  complement  has  been  divided  into  two  parts,  one  present 
in  each  of  the  protein  fractions.  The  globulin  fraction  of  the  com- 
plement will  unite  to  amboceptor  which  is  fixed  to  cells,  and  hence 
is  called  the  mid-piece  of  the  complement,  for  it  will  unite  also  with 

TNofruchi,  Biochem.  Zeit.,  1907   (6),  172. 
TaCourmont  et  nl..  C.  R.  Soc.  Biol.,  1913    (74),   1152. 
7b  Jour.  Infect,  llis.,  1917    (20),  18,5. 

8  Michaelis  and  Skwirsky,  Zeit.  Immunitiit.,  1910   (7),  497. 

1)  Nu<,riichi  and  Bronfonlircniior,  Jour.  K\-i>.  Med.,  19H)  (1.'}),  229;  Rilz,  Zeit. 
Immunitiit.,  1912    (15),  145. 

10  See  Sclimidt,  Arch.  f.  Hy}j.,  1912    (76),  284;  Jour.  llviJ:.,  1914    (14),  4.37. 

11  Zeit.  Immunitiit.,  1913   (10),  503. 


AMBOCEPTOR  AND  COMPLEMENT  213 

the  end-piece  of  tlie  complement  contained  in  the  albumin  fraction, 
and  then  cytolysis  can  take  place.  Without  the  intervention  of  the 
globulin  mid-piece  the  albumin  end-piece  cannot  unite  with  the  am- 
boceptor, while  in  the  absence  of  end-piece  the  amboceptor  mid-piece 
complex  can  cause  no  cytolysis.  Both  fractions  of  the  complement 
are  destroyed  by  heat,  but  if  the  mid-piece  is  bound  to  the  ambo- 
ceptor it  resists  heating.  The  mid-piece  corresponds  to  Ehrlich's 
haptophore,  the  end-piece  to  the  toxophore  group,  and  this  complex 
structure  is  common  to  both  bacteriolytic  and  hemolytic  complement. 
Bronfenbrenner  and  Noguehi,"-''  however,  contend  that  the  supposed 
cleavage  of  complement  is  merely  an  inactivation  by  the  agencies  em- 
ployed, all  the  complement  being  in  the  albumin  fraction  in  a  condi- 
tion capable  of  reactivation,  not  only  by  globulin  but  by  simple 
amphoteric  substances,  a  view  which  has  not  been  generally  accepted. 

In  its  effect  of  dissolving  bacteria  (and  also  other  cells  against 
which  animals  may  have  been  immunized)  complement  resemJjles  the 
enzymes,  and  by  many  it  is  looked  upon  as  related  to.  them,  but  the 
changes  it  produces  do  not  resemble  those  produced  by  proteolytic  en- 
zj-mes  in  all  details."''  In  particular,  complement  seems  to  participate 
in  reactions  according  to  the  law  of  definite  proportions,  unlike  the 
enzymes.^-  In  certain  immune  reactions,  colloids  (lecithin,  silicic 
acid)  ^^  can  play  the  role  of  complement  and  immune  body,  but  these 
reactions  are  probably  quite  different  from  those  of  bacteriolysis  by 
immune  serum. 

Amboceptors  are  formed,  according  to  Wassermann,  and  Pfeift'er 
and  ]\Iarx,  in  the  spleen  and  hemopoietic  organs,  since  in  immuniza- 
tion i\\ey  can  be  demonstrated  in  these  organs  before  they  appear  in 
the  circulating  blood.  The  stability  of  the  amboceptors  is  very  con- 
siderable :  serum  prepared  in  1895  by  Pfeiffer  against  cholera  vib- 
rios was  found  to  have  lost  almost  none  of  its  activity  after  eight 
years  in  an  ice-box  (Friedberger).  Heating  twenty  hours  at  60° 
scarcely  injures  them,  but  70'^  for  one  hour  destroys  them  almost 
completely,  and  heating  the  serum  to  100°  destroj^s  all  the  immune 
bodies.  They  are  quite  resistant  to  putrefaction,  and,  like  the  anti- 
toxins, do  not  dialyze.  Strong  salt  solutions  will  prevent  the  union 
of  complement  and  amboceptor  in  vitro,  and  probably  to  greater  or 
less  degree  in  the  animal  body,  but  the  union  of  antigen  and  ambo- 
ceptor is  not  prevented  by  salt.^*  Alkalies  may  prevent  the  union 
of  amboceptor  with  the  cells,  or  extract  it  from  the  cell  to  w^hich  it 
has  united;  and  they  may  also  inhibit  the  union  of  amboceptor  and 

iia  Jour.  Exp.  Med.,  1912   (5),  598;  good  review  of  literature, 
lib  The  curve  of  coiupleraent  action  resembles  that  of  enzvnic  action.      (Tliiele 
and  Embleton.  -Jour.  Patli.  and  Bact...   191,5    (19),  .372.) 

12  See  Liebermann,  Dent,  nietl.  \Yoch.,   1906    (32),  249. 

13  Landsteiner  and  Jagic,  Wien.  klin.  Woch.,  1904  (17).  03:  Miincli.  med. 
Woch.,   1904    (.51),   1185. 

i4Angerer,  Zeit.  Immunitat.,   1909    (4),  243. 


214  CHEMISTRY    OF    THE    IMMUNITY    REACTIONS 

complement.  Amboceptors  are  not  inactivated  by  shaking,  as  is  com- 
plement, but  they  are  destroyed  alike  by  nltraviolet  rays,  and  both 
resist  a;-rays."^ 

According  to  Pfeiffer  and  Proskauer,^^  digestion  of  the  globulin 
precipitate,  in  which  amboceptors  are  carried  down,  does  not  destroy 
their  activity  completely  even  when  all  the  proteins  are  thus  re- 
moved. Removal  of  the  nucleo-albumin  or  nuclein  does  not  remove 
the  amboceptors  from  the  serum.  Immune  serum  kept  three  months 
in  alcohol  yielded  an  extract  with  distilled  water  that  was  rich  in 
immune  bodies,  but  almost  free  from  protein.  Pick,  Rhodain,  and 
Fuhnnann  found  that  immune  bodies  are  precipitated  entirely  in  the 
euglobulin  fraction  of  the  serum  protein.  From  these  experiments 
it  has  been  thought  by  some  that  the  bacteriolytic  amboceptor  is  not 
itself  a  protein,  although  closely  associated  with  the  serum  globu- 
lins.'® 

CYTOTOXINS 

Just  as  precipitins  can  be  obtained  for  proteins  derived  from  other 
sources  than  bacterial  cells,  so  also  upon  immunizing  an  animal 
against  various  types  of  cells  other  than  bacteria,  substances  appear 
in  its  serum  that  exercise  a  destructive  effect  upon  the  type  of  cells 
injected.  In  other  words,  the  reactions  of  animals  to  infection  are 
not  specially  devised  for  combating  bacteria  and  their  products,  but 
can  be  equally  exerted  against  non-bacterial  cells  and  their  products. 
In  the  case  of  soluble  proteins,  as  before  mentioned,  the  antibodies 
show  their  effects  by  precipitating  them,  with  agglutination  of  the  par- 
ticles into  flocculi  and  perhaps  a  subsequent  digestion ;  in  the  case  of 
cells,  whether  bacterial  or  tissue  cells,  the  antibodies  cause  agglutina- 
tion and  loss  or  impairment  of  vitality.  This  injury  may  be  mani- 
fested by  loss  of  motion  in  the  motile  cells  (bacteria,  spermatozoa, 
ciliated  epithelium)  or  by  solution  of  their  contents  (bacteriolysis, 
erythrocytolysis,  leucocytolysis,  etc.),  or  by  cell  death  without  marked 
morphological  alterations  (B.  typhosus,  spermatozoa).  If  we  inject 
red  corpuscles,  leucocytes,  spermatozoa,  renal  epithelium,  or  any  other 
foreign  cell,  the  reaction  is  as  specific  as  it  is  if  we  inject  bacteria,  and 
of  exactly  the  same  nature.  Therefore,  all  that  has  been  said  pre- 
viously concerning  bactericidal  substances  and  agglutinins  can  be 
transposed  to  apply  to  immunity  against  tissue  cells.  As  a  matter  of 
fact,  however,  the  transposition  is  generally  made  in  the  other  direc- 
tion, for  red  corpuscles  are  much  easier  cells  to  study  than  bacteria, 
because  their  hiking  gives  ])ronipt  and   readily  recognized   evidence 

i*aScamcli,  Uiochoni.   Zeit.,   1015    (60).   102. 

in  Cent.  f.  Bakt.,  189G    (19),  101. 

10  Ascoli  found  that  tlie  active  substance  of  antliracidal  seruni,  which  is  not 
an  amhoceptor,  is  contained  in  the  pseu(h>-j,'h'hulin  fraction  of  asses'  serum,  but 
in  poat's  serum  part  is  in  tlie  euf,'h)bulin  fraction.  (IJiochem.  Centr.,  1006  (5), 
458.) 


HEMOLYSIS  215 

tliat  the  toxic  serum  has  brought  about  changes.  Much  of  our  knowl- 
edge of  bactericidal  serum  has  been  obtained  through  studies  of  the 
mechanism  of  erythrocytolysis,  tlie  results  of  which  have  then  been 
applied  to  the  subject  of  bacteriolysis.  Both  on  this  account,  there- 
fore, and  because  solution  of  red  corpuscles  is  of  itself  an  important 
process  in  manj-  intoxications  and  diseases,  the  subject  is  of  great 
theoretical  and  practical  importance. 

HEMOLYSIS  1'   OR  ERYTHROCYTOLYSIS 

In  hemolysis  the  essential  phenouienon  consists  in  tlie  escape  of 
the  hemoglobin  from  the  stroma  of  the  corpuscles  into  the  surround- 
ing fluid.  As  it  is  not  exactly  known  in  what  way  the  stroma  holds 
the  liemoglobin  normally,  whether  purely  physically  or  in  part  chem- 
ically, or  whether  the  stroma  consists  of  a  spongioplasm  or  sac-like 
membranes,  or  both,  the  ultimate  processes  that  permit  the  escape  of 
the  liemoglobin  are  not  finally  solved.  However,  the  agents  by  which 
the  escape  is  brought  about  are  well  known  and  extensively  studied, 
and  they  are  found  to  be  of  extremely  various  natures.  They  may 
be  roughly  classified  as:  (1)  known  physical  and  chemical  agents;  (2) 
unkno^^Ti  constituents  of  blood-serum;  (3)  bacterial  products;  (4) 
certain  vegetable  poisons ;  (5)  snake  venoms. 

HEMOLYSIS  BY  KNOWN  CHEMICAL  AND  PHYSICAL  AGENCIES 

The  Mechanism  of  Hemolysis. — If  distilled  water  is  added  to 
corpuscles  of  any  kind,  osmotic  changes  are  bound  to  occur,  since 
within  the  cells  are  abundant  salts,  soluble  in  water,  which  will  begin 
to  diffuse  outward  in  an  attempt  to  establish  osmotic  equilibrium  be- 
tween the  corpuscles  and  the  surrounding  fluid.,  Converselj^,  water 
enters  the  corpuscles  at  the  same  time,  and  accumulating  there  leads 
to  swelling  until  such  injurv^  has  been  produced  as  permits  the  hemo- 
globin to  escape  and  enter  the  surrounding  fluid.  Before  this  oc- 
curs the  fluid  is  opaque  because  of  the  obstruction  to  light  offered 
by  the  red  cells,  but  on  the  completion  of  hemolysis  the  fluid  becomes 
transparent.  The  stroma  now  settles  to  the  bottom,  while  the  hemo- 
globin diffuses  into  the  fluid,  making-  it  red,  but  perfectly  transparent. 
This  process  has  long  been  known  as  the  "laking"  of  blood,  and  is 
essentially  the  condition  present  in  all  forms  of  hemolysis.  That  the 
hemoglobin  escapes  only  through  injure*  of  the  stroma  and  not 
through  simple  osmotic  diffusion,  is  shown  l)y  the  fact  that  if  salt 
.solution  of  the  same  concentration  as  normal  serum  is  used  instead 
of  distilled  water,  no  such  escape  of  hemoglobin  occurs.  As  hemo- 
globin is  perfectly  soluble  in  salt  solution,  it  should  pass  out  if  it  dif- 

17  Through  usage  this  term  has  been  limited  to  the  solution  of  the  red  cor- 
puscles, which  is  more  accurately  described  by  the  term  erf/throriitolf/is.  For 
bibliography  see  Sachs,  Ererebniss'e  der  Pathol.,' 1002  (7),  714:  1900  (11),  515; 
Kolle  and  Wassermann's  Handbuch.  191.3  (II),  793;  Landsteiner,  Handbuch  d. 
Biochem.,  1909   (II   (1)),  395. 


216  CHEMISTRX    OF    THE    IMMVSITY    REACTIOXS 

fused  as  do  the  salts.  Since  there  is  no  escape  of  hemoglobin  in  such 
a  salt  solution,  it  is  evident  either  that  the  stroma  is  not  permeable 
to  hemoglobin,  or  else  the  hemoglobin  is  in  some  way  attached  to  or 
combined  with  the  stroma.  Again,  if  the  corpuscles  are  placed  in  a 
solution  of  salt  more  concentrated  than  their  own  fluids,  water  es- 
capes and  the  corpuscles  shrink;  as  no  hemoglobin  escapes  with  the 
Avater,  it  is  evident  that  the  stroma  is  not  permeable  to  hemoglobin 
when  intact.  Because  of  the  resemblance  of  the  process  of  hemolysis 
to  the  rupture  of  plant  cells  with  escape  of  their  contents  when  they 
are  placed  in  distilled  water,  it  might  be  assumed  that  hemolysis  is 
largely  a  physical  matter,  but  if  a  red  corpuscle  in  an  isotonic  solu- 
tion is  cut  into  pieces,  the  hemoglobin  does  not  escape,  indicating 
that  its  structure  is  quite  dissimilar  to  that  of  the  simple  vegetable 
cell,  and  that  there  is  some  union  of  stroma  and  of  hemoglobin, 
whether  a  physical  or  a  chemical  union. ^^  ]\I.  H.  Fischer  ^°  interprets 
hemolysis  as  a  separation  of  lipoid-protein  stroma  and  adsorbed  hemo- 
globin, which  process  can  be  duplicated  experimentally  with  a  com- 
bination consisting  of  a  corresponding  solid  hydrophilic  colloid, 
fibrin,  and  a  hydrophobic  colloid  dye,  carmine;  this  artificial  combi- 
nation behaves  exactly  like  a  corpuscle  to  simple  hemolytic  agents.^'"' 

Repeated  alternate  freezing  and  thawing  is  another  physical  means 
of  bringing  on  hemolysis.  Heating  to  62°-64°  C.  causes  hemolysis 
of  mammalian  corpuscles;  in  cold-blooded  animals  this  seems  to  occur 
at  a  slightly  lower  temperature. 

Some  chemical  agents  are  capable  of  liberating  hemoglobin,  even 
when  the  corpuscles  are  in  isotonic  solutions.  The  ordinary  salts 
of  serum,  of  course,  do  not  have  this  property,  but  annnonium  salts 
are  strongly  hemolytic.  The  chemical  agents  that  dissolve  red  cor- 
puscles seem  to  be  those  that  have  the  power  of  penetrating  the 
stroma.  Ammonium  salts  and  urea  penetrate  the  coii^useles  freelj' 
and  cause  hemolysis.  Sugar  and  NaCl  seem  not  to  penetrate  the 
corpuscles,  and  therefore  do  not  produce  hemolysis.  Of  the  perme- 
ating substances,  there  seem  to  be  two  types :  one,  like  urea,  does  not 
produce  hemolysis  when  in  a  solution  of  NaCl  isotonic  with  the  serum ; 
the  other,  like  ammonium  chloride,  is  not  prevented  from  producing 
hemolysis  by  the  presence  of  NaCl.-° 

18  Stewart  (Jour,  of  Physiol.,  1890  (24),  211)  found  that  in  hemolysis  by 
pliysioal  means  or  under  the  influence  of  servuns,  tliere  is  no  marked  increase 
in  the  electrical  cfmductivity,  but  luMnclysis  liy  sapoJiin  and  by  water  causes  an 
increase  of  conductivity,  presumahl.v  liccaiisc  of  llie  escajjc  of  electrolytes;  cor- 
roborated by  A.  ^^'oelfei,  Biocliem.  ,Jour.,  1908  (3),  140;  see  also  IMoorc  and  Roaf, 
ibid.,  p.  55. 

loKolloid  Zeit.,  1909   (5),  14G. 

ifa  Conccrnin<r  the  influence  of  TT-ion  concentration  on  liciiiolvsis  sec  Walbum, 
Biocheni.  Zeit.,   1914    (6.3),  221. 

-0  Ilainbur<,'er,  in  his  book.  "Osmot  isclier  l^ruck  iiiid  loiiciiiclnf.""  reviews  ex- 
haustively tlie  physical  chemistry  of  hemolysis.  The  followiiiLT  is  his  summary 
of  the  ])cinicability  of  red  cor[niscles  by  various  sulistaiiccs : 


IIIJUOLYSIS  217 

All  these  a^'oiits  seem  to  effect  lieiiiolysis  hif  acliiKj  on  the  stroma, 
for  when  the  stroma  of  corpuscles  hardened  in  formalin  has  its  leci- 
thin and  cholesterol  removed  with  ether,  saponin,  a  powerfully  hemo- 
lytic substance,  seems  to  have  no  effect.  The  action  of  saponin  and 
of  many  other  hemolytic  agents  can  be  prevented  by  the  presence  of 
cholesterol  in  excess,  sug<^esting  that  it  is  this  constituent  of  the 
stroma  that  is  affected.-i^  By  studying  heraol3^sis  under  dark  field 
illumination,  Dietrich  --  found  that  in  water  hemolysis  a  diffusion  of 
hemoglobin  takes  place  through  the  corpuscular  substance,  which  is 
not  visibly  altered;  in  serum  hemolysis  there  is  first  a  precipitate 
formed  in  the  outer  layer,  which  swells.  There  is  no  evidence  that  the 
erythrofeytes  contain  proteolytic  enzymes  of  their  own  that  might  dis- 
integrate them.--^ 

The  fact  that  clioloroform,  ether,  bile  salts,  soaps,  and  amyl  alcohol 
will  cause  laking  is  probably  intimately  connected  with  the  fact  that 
lecithin  and  cholesterol,  important  constituents  of  the  stroma,  are 
both  soluble  in  these  substances.-^  Nearly  all  the  non-specific  hemo- 
lytic agents  are  inhibited  to  greater  or  less  degree  by  the  senim,  in 
which  inhibition  both  the  proteins  and  cholesterol  are  concerned.-* 
Cholesterol  also  influences  many  other  immunity  reactions,  inhibit- 
ing some  and  stimulating  others."^  The  resistance  of  the  corpuscles 
to  hemolysis  by  various  agents  differs  greatly  in  disease,  although 
fairly  constant  in  normal  blood,  the  differences  being  caused  in  some 
cases  by  changes  in  the  permeability  of  the  corpuscles,  and  sometimes 
by  changes  in  the  environment  of  the  corpuscle  or  the  presence  of 
protective  substances  in  either  the  corpuscle  or  the  plasma. 

Arseniuretted  hydrogen,  when  inhaled,  causes  intravascular  hemo- 
l3'sis,  and  there  are  many  other  drugs  and  chemicals  with  the  same 
property,  among  which  may  be  mentioned  nitrobenzol,  nitroglycerin 
and  the  nitrites,  guaiacol,  pyrogallol,  acetanilid,  and  numerous  ani- 

Organic  Substances. —  (a)  Impermeable  for  sugars:  namely,  caiic-sugar,  dex- 
trose, lactose,  also  arabit  and  mannit.  (h)  Permcalilc  for  alcoliols,  in  inverse 
proportion  to  the  number  of  livdroxyl  groups  that  they  contain;  also  for  alde- 
hydes (except  paraldehyde),  ketones,  etliers,  esters,  antipyrin,  amides,  urea, 
urethan,  bile  acids  and  their  salts,  (c)  Slightly  permeable  for  neutral  amino- 
acids    (glycocoll,  asparagin,  etc.). 

Inorganic  substances,  not  including  the  salts  of  the  fixed  alkalies,  (r/)  Com- 
pletely  impermeable  for  the  cations  C'a,  Sr,  Ba,  Mg.  (b)  Permeable  for  XH,  ions, 
for  free  acids  and  alkalies. 

21  Ransom,  Deut.  mcd.  Woch.,  1901  (27),  104;  Kobert,  "Sapoiiinsubstanzen."' 
Stuttgart,  1904;  Abderhalden  and  Le  Count,  Zeit.  cxp.  Path.  u.  TIht..  1!)(1.-)  (•_'). 
199.  Noguclii  (Univ.  of  Penn.  Med.  Bull.,  1902  (15),  ?vll )  found  hvithin 
without  this  property. 

22  Verb.  Deut.  Path.  Gesell.,  1908    (12),  202. 
22a  Von  Roques,  Biocliem.  Zeit.,  1914   (64),  1. 

23  See  Koeppe,  Pfliiger's  Arch.,  1903  (99),  33;  Peskind,  Amor.  .Tour.  Plivs.,  1904 
(12),  184:  Moore,  Brit.  Med.  Jour.,  1909   (ii),  684. 

24  See  V.  Eish^r,  Zeit.  exp.  Path.,  1906    (3),  296. 

25  Walbum,  Zeit.  Immunitiit.,  1910  (7),  544;  Dewev  and  Xuzum,  Jour.  Infect. 
Dis.,  1914  (15),  472. 


218  CHEMISTRY    OF    THE    IMMUMTY    REACTIOXS 

line  compounds.  Probably  the  hemolysis  produced  by  autolytic  prod- 
ucts belongs  in  this  category.""  Alcoholic  extracts  of  tissues  are  com- 
monly hemolytic;  these  extracts  when  added  to  serum  take  on  prop- 
erties which  cause  them  to  resemble  closely  hemolytic  complement 
(Noguchi),  and  the  soaps  seem  to  be  the  active  constituents  of  these 
extracts.  AsHg,  although  strongly  hemolytic  in  the  living  body,  does 
not  hemolyze  corpuscles  in  the  test  tube  (Heffter),  and  this  is  true 
of  some  other  poisons,  which  probably  produce  their  effects  through 
tissue  changes.-'  The  bile  acids  and  their  salts  will  also  produce 
hemolysis,  as  seen  in  jaundice.  Sodium  bicarbonate  solutions  of  one 
or  two  per  cent,  are  hemolytic  for  some  varieties  of  corpuscles,  but 
0.1  per  cent.  NaoCO,,  and  NaHCO.,  do  not  cause  hemolysis.  A  study 
of  the  hemolytic  properties  of  one  class  of  lipolytic  hemolytic  agents, 
the  terpenes,  shows  that  their  hemolytic  activity  varies  much  accord- 
ing to  their  physical  properties,  generally  decreasing  directly  with  in- 
crease in  the  solubility  in  water  (Ishizaka).-^^ 

Leucocytes  are  dissolved  by  some  of  these  agents,  particularly  the 
bile  salts,  although  they  are  affected  by  no  means  so  rapidly  or  so 
much  as  are  the  erj^throcytes.  There  seems  to  be  no  relafion  between 
the  erythrolytic  and  leucolytic  powers  of  these  substances.  Water 
causes  swelling,  with  solution  of  the  granules  in  time,  and  the  same 
is  true  of  ammonium-chloride  solutions. 

Various  chemicals  cause  morphological  alterations  in  the  leucocytes, 
and  of  bacterial  products  the  toxins  of  pyocyaneus  and  diphtheria 
seem  to  be  particularly  leucocidal,  causing  a  striking  karyorrhexis 
(Schiirmann).-^ 

HEMOLYSIS  BY  SERUM 

Normal  blood-serum  of  many  animals  causes  hemolysis  to  greater 
or  less  degree  when  mixed  with  red  corpuscles  of  another  species  of 
animal,  ajid  this  property  can  be  greatly  increased  by  immunizing 
the  animal  with  red  corpuscles  in  the  usual  way.  This  hemolysis  oc- 
curs both  in  the  test-tube  and  in  the  body,  in  the  latter  case  causing 
severe  anatomical  changes  or  even  death.  In  all  respects  the  mech- 
anism of  hemolysis  hij  serum  seems  to  he  idcntieal  with  that  of  bac- 
teriolysis. Two  substances  are  concerned,  one  the  amhocfjytor,  which 
resists  heat  and  which  is  increased  by  immunizing ;  -®*  the  other,  com- 
plement, which  is  destroyed  at  55°  and  which  is  present  in  normal 

29  Concerning  hemolvsis  by  alcoliols,  kotoncs.  etc.,  orjiaiiic  acids,  and  essences, 
see  Vaiidevelde,    ]U\U.  S(ic.  cfiiiii.  dc   ]iolj.M(iue,   lilOf)    (li)),  28S. 

27  Friedljerr,'er  and   Brossa,   Zeit.   Imniunitiit.,    1012    (15),  50(). 
2TaArch.  exp.  Path.,   1914    (75),   195. 

28  Cent.  f.  Patliol.,  1!)10   (21),  337. 

28a  In  an  extensive  study  of  the  hemolytic  antibody,  Thiele  and  Embleton 
(Zeit.  Inmnmitiit.,  101,3  (20),  1)  descril)e  its  forination  as  in  several  steps,  at 
first  bcinj^  tliermolabile  and  unitiiifi  with  tlie  corpiisele  only  when  warmed.  They 
also  find  complement  to  have  several  t-omponents.  If  confirmed,  lliesc  oliserva- 
tions  may  explain  some  of  the  discrciiiuicies  between  the  olisei-vations  and  the 
conclusions  of  different  workers,  which    luive  caused   nianv   controversies. 


HEMOJA'l<Ii^  BY  SERUM  219 

serum.  In  this  case  tlie  substances  may  be  referred  to  as  hemolytic 
amboceptors  and  hemolytic  complements. 

In  spite  of  the  availability  of  these  particular  cytolytic  substances 
for  study,  very  little  has  been  learned  of  their  exact  nature  and 
properties.  It  is  known  that  amboceptor  is  combined  with  the  red 
cells  in  a  certain  sense  quantitatively,  a  definite  amount  being  re- 
quired to  saturate  a  given  amount  of  corpuscles  so  that  they  will  all 
be  hemolyzed  when  complement  is  added ;  and  that  this  reaction  is 
complete  in  less  than  fifteen  minutes  at  45°.  What  change  this  addi- 
tion of  amboceptor  brings  about  in  the  corpuscles  is  unknown.  It 
has  also  been  shown  that  at  0°  the  alifinity  between  the  amboceptor 
and  the  corpuscle  is  greater  than  it  is  between  amboceptor  and  com- 
plement, so  that  it  is  possible  at  this  temperature  to  remove  all  the 
amboceptor  from  a  serum  by  treating  it  with  red  corpuscles,  and 
thus  we  can  obtain  complement  free  from  amboceptor.  This  experi- 
ment also  shows  that  the  two  bodies  exist  side  by  side  in  the  serum 
without  combining,  and  that  combination  occurs  only  after  the  ambo- 
ceptor has  become  united  to  the  erythrocyte.  Hemolysis  by  immune 
sera  takes  place  best  in  a  medium  with  a  reaction  corresponding  to 
that  of  the  blood,  acids  being  more  harmful  than  alkalies;  with  un- 
favorable reaction  the  complement  does  not  unite  with  the  ambo- 
ceptor, although  the  latter  unites  with  the  corpuscle.-'-* 

The  Amboceptor. — Amboceptor  is,  as  a  rule,  destroyed  by  heating 
to  70°  or  higher.-""^  Its  place  of  origin  is  unknowai.  ]\Ietchuikoff 
holds  that  it  is  derived  chiefly  from  the  leucocj'tes,  in  support  of  which 
view  is  the  fact  that  leucocytes  dissolve  red  corpuscles  after  ingest- 
ing them ;  however,  other  phagocytic  cells  have  the  same  power,  par- 
ticularly endothelial  cells,  and  it  is  an  open  question  whether  the  in- 
tracellular digestion  of  engulfed  cells  is  the  same  process  as  extracel- 
lular hemolysis;  probably  it  is  not,  for  there  seem  to  be  more  disin- 
tegrative changes  in  intracellular  digestion  than  in  hemolysis.  Qui- 
nan  ^°  found  that  the  diffusible  constituents  of  hemolj^tic  serum 
played  no  role  beyond  that  of  maintaining  osmotic  pressure.  He 
was  unable,  however,  to  localize  the  immune  body  in  anj'  of  the  pro- 
tein constituents,  and  Liebermann  and  Fenyvess}*  ^^  believe  that  they 
have  obtained  the  amboceptor  in  a  protein-free  condition,  in  which 
it  behaves  like  a  weak  acid.  Amboceptors  are  insoluble  in  lipoids  or 
lipoid  solvents  (Meyer), ^-  and  they  move  towards  the  cathode  in  an 
electric  field,  as  do  other  antibodies.^^  The  amboceptor  complement 
reaction  resembles  a  bimolecular  reaction  which  is  accelerated  by  its 

29Michaelis  and  Skwirsky,  Zeit.  ImnumitUt..   1900    (4K  .357. 
'    29a  Ultraviolet    lio^ht    destroys    immune    hemolysin     (Stines    and    Ahelin,    Zeit. 
Immunitiit.,   1914    (20),  598).' 
"  30  Hofmeister's  Beitr.,  1904   (5),  95. 

3i,Jahresber  d.  Immunitat.,  1911   (7),  2. 

32  Ibid.,  1909,  Vol.  3. 

33  Teague  and  Buxton,  Jour.  Exper.  [Med..  1907   (9),  254. 


220  CHEMISTh'Y    OF    THE    IMMUMTY    REACTIOXS 

end  products  (v.  Krogli).^^  jNIany  of  the  effects  of  hemolytic  ambo- 
ceptors can  be  duplicated  with  silicic  acid ;  ^^  and  a  dye,  brilliant 
green,  may  in  minute  quantities  sensitize  corpuscles  so  that  they  are 
hemolyzed  by  very  small  amounts  of  normal  serum,  or  by  lecithin.^'^^ 

The  amboceptors  of  nonnall.y  hemolytic  serum  seem  to  be  no  differ- 
ent from  those  in  immune  serum,  and  amboceptors  of  one  animal  can 
combine  with  complement  furnished  by  the  serum  of  an  entirely  dif- 
ferent animal.  It  is  tlie  amboceptor  alone  that  gives  the  specific  na- 
ture to  the  reaction,  and,  as  is  the  case  with  all  other  innnunizations, 
it  is  very  diificult  to  secure  antibodies  by  immunizing  an  animal  with 
blood  from  another  animal  of  its  own  species,  isohemolysins.  The 
place  of  origin  of  hemolysins  is  unknown,  as  with  other  antibodies, 
but  that  it  is  not  in  the  blood  seems  to  have  been  established  conclu- 
sively by  Hektoen  ajid  Carlson.^"  Immune  hemolysins  cannot  pass 
from  the  mother  to  the  fetus  before  birth,  but  they  can  be  trans- 
mitted through  the  colostrum  (Famulener).^^ 

Although  Ehrlicli  held  that  the  union  between  cell  and  amboceptor 
is  purely  chemical  and  follows  ordinary  chemical  laws,  especially  the 
law  of  multiple  proportions,  Bordet  and  other  French  observers  have 
claimed  that  the  union  between  amboceptor  and  corpuscle  is  physical 
and  not  chemical.^*  Probably  the  union  is  with  the  stroma  rather 
than  with  the  hemoglobin,  and  the  result  of  the  union  is  to  render  the 
stroma  permeable  to  the  hemoglobin,  or  to  separate  the  bonds  that 
unite  the  hemoglobin  to  the  stroma.^^  There  are  grounds  for  believ- 
ing that  the  amboceptor  not  only  binds  the  complement,  but  that  it 
also  produces  changes  in  the  corpuscles  (^luir).  Mathes  *°  contends 
that  red  corpuscles  cannot  be  dissolved  by  hemolytic  serum  or  by  pan- 
creatic juice  until  after  they  have  been  killed ;  as  heated  serum  does 
not  kill  them,  this  is  presumably  done  by  the  complement.  Corpuscles 
that  have  been  killed  can  then  be  dissolved  in  their  own  serum. 
Levene  *^  tried  to  produce  hemolytic  serums  hy  immunizing  with  dif- 

^34Biochcm.  Zeit.,  1909   (22),  132. 

35  Landstoinor  and  Kock.  Zeit.  Immiuiitiit.,   1912    (14),   14. 

35a  Browninij  and  INIackio,  Zeit.  Tminunitat.,   1914    (21),  422. 

30  .Tour.  Jnfpft.  Dis.,  1910    (7),  319. 

^T  Thid.,  1912    (10),  332. 

38  Ban?,'  and  Forssmann  (Hofmeister'a  Boitr.,  1906  (8),  23S)  suofgcst  that  the 
amboceptor  merely  rendei's  the  corpuscle  permeable  for  tlie  comph'ment,  perhaps 
throuffh  action  on  the  lij)oid  meml)rane;  the  complement  then  acts  directly  upon 
some  constituent  of  the  corpuscle,  witliout  the  amboceptor  actinir  as  a  combining 
substance  in  any  way.  They  found  that  the  substance  in  blood  which  stimulates 
the  antibody  formation  in  the  case  of  hemolysin  formation,  is  chemically  soparabl.> 
from  the  substance  in  blood  whicli  unites  with  these  antibodies;  tlierefore.  they 
conclude,  tlie  "recejitors"  of  cells  are  iiot  identical  with  tlH>  antibodies.  (See  con- 
troversy with  l*]lirlich  in  Miinch.  med.  Woch.,  Vols.  oG  and  57.) 

3!> Corpuscles  treated  with  f)smic  acid  will  unite  with  hemolysins  of  diverse 
oripin,  but  when  used  for  immunizinfi  they  engender  no  hemolysins  (Coca:  also 
V.  Szily,  Zeit.  Imminiitiit.,  1909  (3).  451).  ITeatinfr  corpuscle  stroma  alters 
frreatlv  the  reactivity   (Landsteiner  and  Praselv,  ihid.,  1912    (13),  403). 

•to  Miinch.  med.  Woch.,  1902    (49),  8. 

41  Jour.    Med.    Research,    1904    (12),    191. 


THE  COMPLEMENT  221 

ferent  constituents  of  eorpuselcs,  usiii<^ — (1)  pure  crystalline  hemo- 
globin; (2)  proteins  of  the  stroma  soluble  in  salt  solutions;  (3)  an 
extract  with  alcohol-ether;  and  (4)  an  extract  in  1.5  per  cent,  sodium 
bicarbonate.  Only  the  last  gave  positive  results,  and  the  serum  was 
almost  devoid  of  agglutinative  ])roperties.  Injection  with  corpuscles 
that  liad  been  digested  witli  trypsin  gave  about  the  same  results  as 
alkaline  extracts;  corpuscles  digested  by  pepsin  gave  a  much  weaker 
serum ;  in  neither  was  agglutination  obtained.  According  to  Bang 
and  Forssmann  '-  ethereal  extracts  of  red  corpuscles  give  rise  to  pro- 
duction of  hemolysins  on  imnumization,  and  this  "lysinogen"  sub- 
stance can  be  precipitated  with  acetone,  is  insoluble  in  alcohol,  is  not 
destroyed  by  boiling,  and  gives  rise  to  no  agglutinin.  Ford  and  Hal- 
sey  *^  obtained  serum  with  both  lytic  and  agglutinative  powers  by  in- 
jecting either  the  stroma  or  the  laked  blood  free  of  stroma ;  results 
with  pure  hemoglobin  were  indefinite.  Stewart  *^  obtained  similar  re- 
sults by  immunizing  with  corpuscles  laked  by  physical  means,  by 
serums,  or  by  saponin.  According  to  Guerrini,*^  nucleoprotein  ob- 
tained from  dog's  blood  will  give  rise  to  specific  hemolysins,  and  Beebe 
states  that  nucleoproteins  from  visceral  organs  do  not  have  this  effect. 
Levene's  alkaline  extracts  probably  also  contained  nucleoproteins. 
Immunization  with  extracts  of  tissues  and  cells  of  various  sorts,  even 
when  entirely  free  from  blood  (e.  g.,  spermatozoa),  may  produce 
hemolytic  sera.  The  fact  that  various  tissues  from  many  different 
species  of  animals,  when  used  as  antigen,  may  give  rise  to  hemolysin 
for  sheep  corpuscles,  is  an  interesting  but  so  far  unexplained  phe- 
nomenon, which  is  discussed  under  "Specificity"  in  the  preceding 
chapter. 

The  Complement. — Hemolj'tic  complement  possesses  the  same 
properties  as  bacteriolytic  complement,  resembling  enzymes  to  the  ex- 
tent that  it  is  susceptible  to  heat  and  causes  a  disintegration  of  cells, 
and  is  largely  retained  by  Berkefeld  filters.*"  The  joint  action  of 
amboceptor  and  complement  is  strikingly  like  the  activation  of  tryp- 
sinogen  by  kinase.  On  the  other  hand,  hemolysis  by  serum  is  quite 
different  from  the  effect  of  trypsin  on  corpuscles,  as  trypsin  completely 
disorganizes  the  hemoglobin  and  destroys  the  stroma,  while  in  hemo- 
lysis the  stroma  and  hemoglobin  seem  to  be  merely  separated  from  one 
another  but  not  chemieallj'  altered.  Again,  hemolysin  acts  quanti- 
tatively, although  that  may  be  due  to  a  difference  in  the  way  the  bind- 
ing to  the  cell  occurs,  rather  than  in  the  method  of  action  of  the  com- 
plement. Landsteiner  and  others  have  suggested  that  a  lipoidal 
complement  dissolves  the  corpuscle  lipoids,  liberating  the  hemoglobin, 
while  Neuberg  and  others  haA^e  supported  the  hypothesis  that  comple- 

42  Hofmeister's  Beitr.,  1906   (8),  238. 

—  43  Jour.  Med.  Research.   1004    (11),  40.3. 

-  44Amer.  Jour,  of  Phvsiol.,  1004    (11).  250. 
45Puv.  crit.  di  din.  liied..  1003    (4),  561. 

4GMuir  and  Browning,  Jour.  Path,  and  Bact.,   1009    (13),   232. 


222  VUKMfsTh'Y    OF    THE    JMMi.MTY    L'EACTIOSS 

ineut  is  virtually  a  lipase  which  splits  the  lipoids  out  of  the  corpuscles. 
Bordet  believes  that  the  hemolysin  causes  a  lesion  of  the  stroma  which 
changes  the  resistance  to  osmotic  influences.  Dick'*^  has  found  evi- 
dence that  the  complement  is  a  ferment  formed  in  the  liver,  and 
that  it  causes  actual  proteolytic  changes.  Jobliug  ^^  associates  the 
serum  lipase  with  the  hemolytic  complement.^*"'  Ohta'"'  observed  no 
increase  in  non-coagulable  nitrogen  during  hemolj'sis,  but  Dick 
found  an  increase  in  the  free  amino  acids ;  therefore,  as  yet  agreement 
has  not  been  reached  as  to  whether  hemolysis  depends  in  any  way 
upon  proteolysis  or  lipolysis  in  the  corpuscle  stroma. 

Although  the  serum  of  one  animal  may  complement  the  immune 
bodies  in  serum  of  several  other  varieties,  and  also  produce  lysis  of 
many  sort  of  cells,  it  may  be  that  not  one  complement  does  all  the 
complementing ;  Elirlich  and  others  have  asserted  tliat  one  serum  may 
contain  several  complements  of  slightly  differing  natures.  Noguchi,°° 
Liebermann  and  Fenyvessy,  and  others  have  pointed  out  the  striking 
resemblance  between  hemolytic  complement  and  certain  compounds  of 
soaps  or  lipoids  with  serum  proteins,  and  it  is  possible  that  such 
compounds  are  of  importance  in  serum  hemolysis;  but  there  seems 
also  to  be  evidence  of  the  existence  of  distinct  protein  complements, 
entirely  different  from  these,^^  and  it  is  possible  that  the  protein  com- 
plements are  the  important  agents  in  specific  hemolysis  by  immune 
sera." 

Antibodies  can  be  obtained  for  both  complement  and  hemolytic 
amboceptor  by  immunizing  against  serum  containing  them,  and  in 
many  serums  antihcmolysins  exist  normally.  Against  certain  vegeta- 
ble hemolysins  this  antihemolytic  action  is  \evy  strong  (Kobert). 
Antihcmolysins  are  generally  anticomplements,  but  in  a  number  of 
instances  anti-amboceptors  have  been  obtained.  The  existence  of  im- 
mune bodies  specific  for  hemolytic  amboceptor  and  complement,  sup- 
ports the  view  that  both  of  these  agents  are  proteins. 

Hemagglutinin. — Agglutination  of  red  corpuscles  occurs  under  the 
influence  of  immune  serum  as  well  as  under  the  influence  of  some 
normal  serums.     In  all  respects  the  principles  seem  to  be  the  same  as 

47  .Tour.  Infect.  Dis..  1913    (12),  111. 

•ts.Toblinfr  and  Bull,  .Tour.  Exper.  Med.,  101.3  (17),  (51:  also  Bergel.  Deut.  Arch, 
klin.  Med.,  1012   (100),  47. 

48a  Thiele  and  Embleton,   however,   state  that  hemolysin   is  not  a   lipase,   and 
that  tlie  hemohtic  power  of  serum  has  no  relation  to  its  lipolytic  power    (Jour. 
Path,  and  Bact.,   1914    ( 10) ,  349) . 
"49Biochem.  Zeit.,  1012   (40),  247. 

soBiochem.  Zeit.,  1907    (6),  172  and  327;   Jour.  Exper.  Med.,  1907    (0),  430. 

•'■1  See  Liefmann.  et  al.,  Zeit.  Imnuuiitiit.,  1012   (13),  150. 

52  Liebermann  and  Fenyvessy  (loc.  cit.)  si  ludieve  that  serum  hemolysis  takes 
place  as  follows:  First,  tlie  aml)0('eptor  acts  on  the  corpust'Ic,  injurinL;  it  so 
that  it  becomes  h-ss  resistant;  second,  tliis  combination  acts  upon  the  comple- 
ment (a  soap  compound)  and  frees  the  soap  so  tliat  it  can  unite  with  tlie  ambo- 
ceptor-corpuscle  system;  third,  tlie  soaj)  causes  liemolysis;  fourth  (aa  a  separate 
step),  the  escape  of  the  hcnio^'h)])iii   from   tlie  corpuscles. 


llEMAdCIJ  TIMN  223 

those  described  for  bacterial  aygiutiiiatioji.  Tlie  hemagglutinating 
antibody  behaves  like  the  other  antibodies  and  proteins  under  the  in- 
fluence of  chemical  and  physical  agencies,  but  Landsteiner  and  Jagic 
have  obtained  strong  agglutinating  solutions  containing  very  little 
protein.  Bergel  '^  contends  that  hemagglutination  is  produced  by 
lipase  from  the  lymphocytes,  which  alters  the  lipoid  membranes  of 
the  erythrocytes.  Agglutination  occurs  at  much  lower  temperatures 
than  hemolysis,  and  also  is  not  checked  h\  heating  the  serum  to  55° ; 
hence  it  is  possible  to  observe  hemagglutination  independent  of  hemo- 
lysis. Serums  may  contain  hemagglutinins  and  not  "be  hemolytic; 
the  reverse  is  also  true.  The  conglutinin  effect  of  beef  serum  (Bordet 
and  Gay)  is  also  observed  with  corpuscles  as  with  bacteria.  As  agglu- 
tination occurs  in  corpuscles  that  have  been  fixed  in  formalin  or  sub- 
linuite,  it  is  probably  not  the  proteins  that  are  affected,  but  some  other 
of  the  ingredients  of  the  stroma,  of  which  lecithin  and  cholesterol  seem 
to  be  the  chief. 

Certain  vegetable  poisons  also  produce  agglutination  of  red  cor- 
puscles, especially  ricin,  abrin,  and  crotin,  and  the  fact  that  ricin 
has  little  or  no  hemolytic  action  shows  the  independence  of  the  proc- 
esses. Antisera  for  these  vegetable  poisons  are  also  antiaggiutina- 
tive,  acting,  as  Ehrlich  showed,  on  the  poison  and  not  on  the  corpus- 
cles. The  seeds  of  many  non-poisonous  leguminous  plants,  and  also 
of  Solanacece,  jdeld  extracts  that  are  strongly  agglutinative  for  red 
corpuscles;  in  Phaseolus  multiflorus  the  active  substance  is  found 
in  the  proteose  of  the  seed,  and  seems  to  be  a  part  of  the  stored  food 
(Schneider).^*  It  is  not  present  in  other  parts  of  the  plant.  Snake 
venoms  contain  agglutinins,  destroyed  by  heating  to  75°  ;  their  ag- 
g-lutinating  power  being  in  inverse  ratio  to  their  hemolytic  power. 
Corpuscles  agglutinated  by  venoms  may  be  again  separated  by  po- 
tassium permanganate  solutions.^^  Silicic  acid  and  certain  other 
colloids  may  act  as  agglutinins,  their  effects  bearing  a  relation  to  the 
effects  of  electrical  charges  upon  agglutination  of  bacteria  or  of  col- 
loids {q.  v.).-'^  Corpuscles  that  have  been  sensitized  by  hemolytic  am- 
boceptors are  much  more  readily  agglutinated  by  salts  of  heavy  metals, 
especiall}'^  copper  and  zinc,  presumably  because  of  quantitative  altera- 
tions in  the  electrical  charge  of  the  corpuscles  induced  by  the  anti- 
body.""'"^ 

Agglutination  of  the  corpuscles  during  life  va&y  be  of  great  patho- 
logical importance,  for  such  masses  of  agglutinated  corpuscles  may 
readily  produce  capillary  thrombi  and  emboli,  which,  if  wide-spread, 
may  create  much  disturbance.  Sometimes  the  serum  of  one  indi- 
vidual of  a  species  agglutinates  the  corpuscles  of  another  individual 

53Zeit.  Immunitat.,  1912    (14),  255;    1913    (17),  169. 

54  Jour.   Biol.   Chem.,    1912    (11),   47;    bibliographv. 

55  See  Flexner,  Univ.  of  Penn.  Med.  Bull.,  1902   (1.5).  .*324  and  301. 

56  See  Landsteiner  and  Jagic,  Miinoh.  med.  Woch..   1904    (51),   1185. 
56a  Eisner  and  Friedemann,   Zeit.   Immunitiit.,   1914    (21),  520. 


224  CHEMISTRY    OF    TIIK    IMMUMTY    REACTIONS 

of  the  same  species  {isoayglutination) ,  a  fact  which  must  be  taken 
into  account  in  performing"  transfusion  of  blood,  lest  dangerous  ag- 
glutination take  place.  Agglutination  of  an  individual's  corpuscles 
by  his  own  seinim  (autuaggliitination) ,  nuiy  also  be  observed  under 
experimental,  and  perhaps  under  pathological  conditions  (Land- 
steiuer),  this  pathological  autoagglutination  probably  occurring  espe- 
cially at  temperatures  below  37°.  (See  Parox.ysmal  Hemoglobin- 
uria.) ]\Iany  bacteria  produce  substances  that  are  agglutinative  for 
human  red  corpuscles,  among  them  being  B.  typhosus,  pyocyaneus, 
and  staphylococcus.  Flexner  ^^~  has  found  in  typhoid  fever  thrombi 
that  seemed  to  be  composed  of  agglutinated  red  corpuscles,  almost 
free  from  fibrin  and  leucocytes.  Probably  many  of  the  so-called  "hy- 
aline thrombi"  found  freciuently  in  infectious  diseases  are  really  com- 
posed of  agglutinated,  partly  hemolyzed  red  corpuscles  (see  "Throm- 
bosis," Chap.  xi). 

HEMOLYSIS  BY  BACTERIA -s 

Both  pathogenic  and  non-pathogenic  bacteria  produce  hemolytic 
substances  that  are  excreted  into  the  fluids  in  which  they  grow.  Dur- 
ing many  infectious  diseases  marked  hemolysis  occurs,  especially  in 
those  diseases  accompanied  by  septicemia.  After  death  the  hemo- 
globin of  the  blood  goes  into  solution,  and  the  resulting  staining  of 
the  walls  of  the  blood-vessels,  and  later  of  the  tissues  everywhere,  is 
generally  familiar.  In  the  post-mortem  hemolysis  probably  the  pu- 
trefactive organisms  are  chiefly  concerned,  although  it  is  marked  a 
very  short  time  after  death  in  many  cases  of  septicemia,  particularly 
when  the  infecting  organism  is  the  streptococcus,  and  here  probably 
the  pathogenic  organism  is  the  chief  cause  of  the  hemolysis.  The 
hemolytic  action  of  bacteria  can  be  studied  both  {}i  vitro  and  in  vii'o. 
Among  the  best  known  hemolytic  bacterial  toxins  are  tetaiwlysin, 
pyocyanolysin,  typholysin,  staphylolysin,^^  and  streptocolysin,  as  they 
have  been  termed.  Of  these,  the  case  of  pyocyanolysin  is  question- 
able, because  it  has  been  described  as  resisting  heat  above  the  boiling- 
point,  and  Jordan  *"'  seems  to  have  proved  that  the  hemolysis  is  a.scriba- 
ble  to  the  alkalinity  that  this  organism  produces  in  culture-media. 
Other  bacterial  hemolysins  are,  however,  destroyed  by  heat  at  70°  or 
less  for  two  hours ;  but  they  are  altogether  different  from  ordinary 
cellular  hemolysins.     Apparently  streptocolysin  is  simply  a  toxin  for 

57  Univ.  of  R-nn.  IMod.  Bull.,  1002  (15),  324';  Aini-r.  Jour.  Mod.  Sci.,  I'M);!  (  12(i), 
202. 

58  See  Pribram,  Kolle  and  Wassermann's  Handliuch..   lOi;^    (II).   l:?2S. 

59  Analysis  of  staphylolysin  ])v  Burkhardt  (Arcli.  cxp.  I'itth.  \uul  IMiarni..  I!tl0 
(63),  107),  showed  it  to  1)p  diiily/.ahlc,  protein-  and  biiiiot-ficc.  tlicrniolaliilo  and 
soluble  in  other.  From  1i.  piitidum  lie  isolated  a  luMnolytic  substance  which 
seems  to  be  a  derivative  by  oxidation  of  erucacic  acid  ( oxydiiut'lliylthiolerucacic 
acid ) . 

'^I'.lour.  Medical  Research,  100:]   (10),  31. 


HEMOLYSIS  BY  YEGETABIJ-:  POISONS  225 

red  cells,"'  and  unites  directly  to  the  cell  receptors  without  the  inter- 
vention of  any  intermediary  body.  As  a  similar  structure  has  been 
shown  for  stai)hylolysin  and  tetanolysin,  it  is  probable  that  the  hac- 
ierial  hemolysins  are  all  merely  toxins  tcith  a  particular  affinity  for 
red  cells,  and  ajrainst  some  of  these  bacterial  hemotoxins  antitoxic  sera 
are  obtainable,  althouo-h  there  is  usually  some  question  as  to  how  much 
of  tlu:"  antagonistic  effect  depends  on  true  antitoxins  and  how  much 
upon  the  cholesterol  in  the  scrum.  Of  course  bacteria  may  also  fonii 
many  non-specific  hemolytic  sid)stances  as  products  of  their  metab- 
olism, such  as  acids  and  bases. 

Secondary  anemia  occurring  in  the  infectious  diseases  is  probably 
to  be  explained  largely  by  this  hemolj'tic  property  of  bacterial  toxins. 
Hemoglobinuria  may  also  be  produced  in  the  same  way  in  some  in- 
stances. Intravenous  injections  of  filtrates  of  the  saprophyte,  B. 
megatherinm,  will  produce  hemoglobinuria  in  guinea-pigs,  hence 
hemolysis  is  not  an  exclusive  property  of  pathogenic  bacteria,  and  with 
streptococci  Lyall  '^^^  found  that  the  hemolysin  titer  did  not  afford  a 
criterion  of  virulence.  No  immunity  is  produced  in  animals  immun- 
ized with  streptococcus  hemolysin."'^  Pneumoeocci  produce  an  intra- 
cellular hemolytic  toxin  which  is  very  labile  and  antigenic ;  living 
pneumoeocci  convert  hemoglobin  into  methemogiobin,  but  this  the 
hemolytic  extracts  of  pneumoeocci  cannot  do  (Cole).^^'^  Streptococcus 
viridans  has  the  same  property,*^'*  which  may  play  a  part  in  the  effects 
of  infections  with  these  organisms. 

HEMOLYSIS  BY  VEGETABLE  POISONS 

A  nuudjer  of  plant  poisons  are  strongly  hemolytic,  and  some  of 
them  owe  much  of  their  toxicity  to  their  effect  on  the  erythroc^'tes. 
One  group  consists  of  the  bodies  often  called  "vegetable  toxalbu- 
mins, "  because  they  seem  to  be  proteins,  and  includes  ricin,  abrin, 
crotin,  curcin  and  robin.  Of  these,  crotin  and  curcin  are  particularly 
actively  hemolytic,  while  ricin,  abrin,  and  robin  are  more  marked 
by  their  agglutinating  action,  hemolysis  being  produced  only  by 
relatively  large  doses.  Their  effects  varv^  greatly,  however,  according 
to  the  species  of  animals  whose  blood  is  used.  They  resemble  the 
bacterial  toxins,  in  that  immunity  can  be  secured  against  them,  and 
the  imnume  serum  will  prevent  their  hemolytic  action.  Heating  the 
toxalbumins  to  65°  or  70°  does  not  destroy  the  hemolytic  or  agglu- 
tinating action  except  with  phallin,  but  100°  does.  The  action  of 
these  substances  is  not  like  that  of  the  enzymes,  in  that  it  is  quanti- 
tative,, a  given  amount  acting  on  a  given  amount  of  corpuscles  to 

61  Jour.  Amer.  Med.  Assoc,  1003    (41),  0G2;  Jour.  Infect.  Dis.,  1007    (4),  277. 

cia.Jour.  Med.  Res.,  1914   (30),  51.5. 

sibMcLeod  and  McXeo.  .Tour.  Path,  and  Bact.,   1013    (17),  524. 
->   sicJour.  Exper.  ^led..  1914   (20),  347.  3(!3. 
-   eid  Blake,  Jour.  Exper.  Med.,  1916    (24),  315. 
15 


226  CHE^[IsTl,•y  or  rni:  immimtv  reactioxs 

which  it  is  bound.  ^Madsen  and  Walbuni  '^-  observed  that  red  corpiis- 
t'les  had  the  power  of  dissociatino-  neutral  mixtures  of  ricin  and  anti- 
riein,  the  ricin  entering  the  corpuscles  from  which  it  could  be  recov- 
ered.®^ Ford  and  Abel  believe  the  hemolj^tic  agent  of  amanita  to  be 
a  glucoside.  (The  general  nature  and  other  properties  of  these  sub- 
stances are  considered  under  the  heading  of  "Phytotoxins, "  in 
Chap,  vi.) 

Saponin  Group. — Another  quite  distinct  group  of  vegetable 
hemolyzing  agents  consists  of  the  ^'saponin  substances."  ^^  These 
are  a  closely  related  group  of  glucosides,  found  in  at  lea.st  46  differ- 
ent families  of  plants,  and  they  are  strong  protoplasmic  as  well  as 
hemolytic  poisons.  They  differ  altogether  from  the  true  toxins,  be- 
ing heat  resistant,  having  no  resemblance  to  proteins,  and  not  giving 
rise  to  antibodies  on  immunization  of  animals. "^^  The  degree  of  their 
toxicity  is  not  directly  proportional  to  their  hemolytic  activity ;  they 
seem  to  injure  chiefly  the  nerve-cells.  Apparently  hemolysis  is 
brought  about  by  action  upon  the  lipoids  of  the  red  corpuscles,  for 
addition  of  cholesterol  to  saponin  prevents  its  hemolytic  effect ;  "*'  leci- 
thin does  not  have  the  same  property.®^  Both  cholesterol  and  leci- 
thin combine  M'ith  saponin,  the  cholesterol  compound  being  quite 
inert,  whereas  the  lecithin  compound  is  both  hemolytic  and  toxic. 
The  compound  formed  between  a  typical  saponin,  digitonin,  and 
cholesterol,  is  so  insoluble  that  it  has  been  found  useful  in  the  quan- 
titative analysis  of  cholesterol."^  Normal  serum  seems  to  contain 
an  antihemolysin  for  saponin,  and  therefore  hemoglobinuria  is  not 
produced  by  all  saponins  on  intravenous  injection.  Careful  immu- 
nization leads  to  a  slight  increase  in  this  antihemolytic  action  of  the 
serum,  possibly  due  to  an  increased  formation  of  cholesterol  (Ro- 
bert). The  resistance  of  corpuscles  to  saponin  hemolysis  varies  in 
disease,  being  especially  low  in  jaundice  (M'Neil).*"' 

A  study  of  the  toxicity  of  the  members  of  this  group  by  Kobert '° 
shows  that  in  general  they  have  similar  properties,  but  that  minor 

c2Cent.  f.  Bakt.,  in04    (36).  242. 

IS  According  to  Pascueoi  (Hofmeister's  Bcitr..  1905  (7).  4.5").  riein  combines 
directly  with   lecitliin,  the  compound  beinjr  strongly  liemolytic. 

f*  Completo  litoraturc  on  saponin  pivon  by  Kobert.  "Die  Raponinsnbstanzen." 
Stuttgart,  1004:   also  Kuiikel.  "Handi)iicli  der  'I'oxokolotrie,"  .Tena. 

6'''  Saponins  are  cliaracterized  by  tlieir  ready  solubility  in  water  and  the 
foamino:,  soapy  character  possessed  by  tlie  solution;  hence  their  teclinical  appli- 
cations as  soap  bark,  etc.  Heated  with  dilute  acids  they  split  ofT  sugar;  also 
when  acted  on  by  frlucoside-spliUin"r  enzymes  (from  spiders),  accordinsr  to 
Kobert.  Saponin  from  QiiiUnja  (soajj-bark)  has  the  formula  f",„lT;,„0,o  (Stiitz). 
Most  are  colloids,  but  some  crystallize. 

66  Ransom,  Dent.  mcd.  Woch.,  1001  (27).  104:  INbidsen  and  Xounulii.  Cent.  f. 
Bakt..  100.5    (.'^7).  .367;  Pascucci,  Hofmeister's  Beitr..  1005    (6).  .543. 

OTXopuchi,  Univ.  of  Benn.  Med.  Bull.,  1002  (15),  .327:  Meyer.  Hofmeister's 
Beitr..   1008    (11),  357. 

«8Windaus,   C'hem.   Berichte.    1000    (42),  238. 

co.Tour.  Path,  and  Bact..  1010   (15),  56. 

70  Arch.  exp.  Path.  u.  Pharm.,  1887    (23),  233. 


THE  ^AI'OM.y  a  ROUP 


227 


differences  exist  between  tlieni.  All  cause  hemolysis,  some  in  .Illa- 
tion as  great  as  1 :100,000.  Some  produce  hemoglobinuria  when  in- 
iected  intravenously,  others  do  not.  All  paralyze  the  heart,  but  the 
injuries  to  the  central  nervous  system  are  the  chief  cause  of  death, 
^larked  local  changes  are  produced  at  the  site  of  injection,  but  the 
leucocytes  are  apparently  not  injured,  although  sterile  suppuration 
is  produced.  There  is  a  period  of  latency  after  intravenous  injection 
of  small  doses— twenty-four  hours  or  more— before  the  appearance 

of  symptoms. 

Sapotoxin  is  one  of  the  most  actively  toxic  and  hemolytic  products 

of  quiUaja.  .      -,  ^         ^     ^  \ 

Cyclamix  is  also  a  member  of  this  group  (derived  from  Cyclamen), 
and  is  said  to  be  the  most  active  of  all  as  a  hemolytic  agent  (Tufa- 

now).  ,     ^  1  •     J 

SoLANix  '^  is  obtained  from  all  parts  of  the  potato  plant,  combined 
with  malic  acid;  it  is  found  particularly  in  young  sprouts,  but  not 
in  any  considerable  amounts  in  normal  potatoes.^^  Its  formula  is 
unknown  but  as  it  splits  up  into  an  alkaloid  (solanidin)  and  sugar 
it  is  called  a  glyco-alkaloid.  In  its  action  it  resembles  the  saponins, 
being  a  powerful  protoplasmic  poison,  killing  bacteria,  and  hemolyz- 
ing  blood  in  very  great  dilutions. 

A  great  number  of  hemolytic  poisons  are  obtained  from  poisonous 
mushrooms.     Best  known  of  these  is : 

Helvellic  Acid,  from  HelveUa  esculenta,  which  has  the  empiric 
formula  Ci,H„oO,."  Intravenously  injected  it  produces  hemoglobin- 
uria  and   icterus,   with   hemoglobin   infarcts   in   the   kidneys    (Bos- 

troem).'^*  , 

Phallix,  or  Amanita  hemolysin,  described  by  Robert  as  a  toxal- 
bumin  has  been  found  by  Abel  and  Ford  to  be  a  glucoside,  and  thus 
belongs  to  the  saponin  group.  (See  Chap.  vi.  for  further  discussion.) 
In  the  leaves  of  the  ivy,  Hcdera  helix,  a  hemolytic  glucoside  has  been 
found  by  Moore.''  It"  is  of  interest  that  Faust  believes  the  poisonous 
agent  of  cobra  venom  to  be  a  glucoside,  closely  resembling  sapo- 
toxin. 

As  will  be  seen,  all  these  last-mentioned  vegetable  hemolytic  agents 
are  essentially  different  from  either  the  bacterial  or  serum  hemolysins, 
or  from  the  abrin,  ricin,  crotin,  or  robin  group,  in  that  they  are  of 
relatively  simple  chemical  composition,  and  quite  unlike  proteins,  en- 
zymes, or  toxins.  The  manner  in  wdiich  they  cause  hemolysis  is 
unknown,  but  from  their  relation  to  saponin  it  is  probable  that,  like 
it,  they  cause  injury-  by  combining  with  or  dissolving  the  lipoids  of 

Ti  Literature,  see  Mever  and  Schmiedeberg,  Arch.  f.  exp.  Path.  u.  rharm.,  1895 
(36).  361:   Perles,  ibid.,  1890    (26),  88. 

72  See  Kunkel,  "Handbueh  der  Toxokoloie,"  p.  873. 

73Boehm  and  Kiilz,  Arch.  exp.  Path.  u.  Pharm.,  1885    (19),  403. 

74Deut.  Arch.  klin.  Med.,  1883   (32),  209. 

T5  Jour.  Pharmacol.,  1913   (4).  263. 


228  CHEMLSTRY    OF    rilE    IMMl  MTV    REACTIONS 

the  stroma  of  the  corpuscles.  Extracts  of  Morchella  esculcnta  do  not 
hemolyze  corpuscles  in  vitro,  although  powerfull}^  hemolytic  when 
injected  into  animals,  and  causing  severe  hemoglobinuria;  so  that 
it  is  probable  that  they  cause  their  hemolytic  effects  indirectly  through 
the  changes  which  they  produce  in  the  tissues  of  the  poisoned  animal.'^'' 

HEMOLYSIS  BY  VENOMS" 
The  laking  of  blood-corpuscles  by  venoms  is  of  peculiar  interest 
from  the  standpoint  of  immunity  phenomena,  since  it  was  demon- 
strated by  Flexner  and  Noguchi  that  the  hemolytic  principle  of  the 
venoms  resembles  an  amboceptor,  in  that  some  substance  behaving 
like  complement  has  to  be  furnished  by  the  blood.  Kyes  demon- 
strated that  this  complementing  agent  is  lecithin,^^''  and  was  able  to 
produce  what  he  considers  to  be  compounds  of  the  hemolysin  with 
lecithin,  called  "leeithids."  The  hemolytic  activity  of  these  lecithids 
is  very  great,  and  they  seem  to  be  free  from  the  neurotoxic  princi- 
ple of  the  venoms.  Whether  thej^  represent  true  compounds  of  a 
hemolytic  amboceptor  with  lecithin,  or  are  simpl}^  actively  hemolytic 
products  of  the  cleavage  of  lecithin  by  an  enzymatic  activity  of  the 
venom,  is  at  present  unsettled ;  ^^  it  seems  probable,  however,  that 
the  hemolysin  of  cobra  venom  is  a  lipase  that  splits  lecithin  into  two 
hemolytic  components,  oleic  acid  and  "desoleolecithin"  (Coca)." 
Noguchi  suggests  that  not  only  lecithin,  l)ut  also  soaps,  especially 
of  unsaturated  fatty  acids,  and  probably  protein  compounds  of  soaps 
jind  lecithin,  may  act  as  the  hemolytic  "complement"  which  activates 
venoms.  The  hemolytic  agents  of  venom  seem  to  be  secreted  by  the 
salivary  glands  of  the  reptiles  from  their  blood,  wliieh  contains  almost 
identical  amboceptors,  differing  chiefly  in  that  they  can  be  activated 
only  by  agents  contained  in  snake  blood,  while  the  amboceptors  of 
venom  can  be  activated  by  nearly  all  sorts  of  blood.  Venoms  from 
cobra,  rattlesnake,  moccasin,  and  copperhead  possess  in  each  a  variety 
of  intermediary  bodies  (amboceptors)  that  seem  to  be  at  least  partly 
identical  in  nature,  although  they  may  vary  in  quantity.  In  order  of 
decreasing  hemolytic  power  for  mammalian  corpuscles  come  venoms 
from  cobra,  water  moccasin,  copperhead,  and  rattlesnake.  These 
venoms  are  also  agglutinative  for  all  corpuscles  tried,  and  agglutina- 
tion will  occur  at  0°  C.  Exposure  for  thirty  minutes  at  75°-80°  C. 
destroys  the  agglutinating  property.  In  general,  the  hemolytic  power 
of  the  venoms  for  different  sorts  of  corpuscles  varies  in  inverse  ]iro]ior- 

70  Freidborcrer  and  Brossa,  Zoit.  Immunitiit.,   1912    (15),  .506. 

77  Geneial  review  of  literature  on  the  lieniolytie  jirojjerties  of  animal  poisons 
pivcn  by  Saelis,  r.ioelieiii.  Centrallilatt,  IHOG  (.ij.  257;  No-ruclii,  Jour.  Kxp.  Med., 
1907   (!)).  4:m. 

77a  Cruicksliank  also  found  tliat.  oilier  ]i])oids  tlian  lecitliin  niav  activate  eobra 
venom    (Jour.  Patli.  and  IJaet.,   1013    (17),   019). 

78  See  Kyea,  Jour.  Infeet.  Dis.,  1910  (7),  181:  v.  Dun,u:erii  and  Coea.  ihiiJ..  1912 
(10),  57 ;  "Manwarinj:.  Zeit.  Immunitiit.,  1910  ((5),  r)i:{;  I?aii;r,  ihid..  I'.Hi)  (S), 
202;  Coca,  Jour.  Infect.  Dis.,  1915   (17),  351. 


HEMOLYSIS  IX  DISEASE  229 

tion  to  their  agglutinative  power.  The  hemolytic  iutermediary  bodies 
are  resistant  to  heat,  suffering  but  slight  loss  of  power  at  100°  C. 
•Red  corpuscles  of  the  frog  are  not  hemolyzed  by  venom,  and  those  of 
■necturus  (mud  pui)py )  but  slightly,  agreeing  with  the  known  resist- 
ance of  cold-blooded  animals  to  snake-bites. 

The  erythrocytes  of  different  individuals  show  considerable  varia- 
tions in  their  resistance  to  hemolytic  agents,  perhaps  depending  upon 
the  amount  or  upon  the  manner  of  fixation  of  the  lipoids  in  the  cor- 
puscles; thus,  the  corpuscles  of  syphilitics  show  a  heightened  resist- 
ance to  hemolysis  by  cobra  venom  (Weil)  "  except  in  the  earliest 
stages,  when  they  are  hypersensitive.  Also,  the  serum  of  persons  suf- 
fering from  various  diseases,  especially  mental  diseases,  inhibits  the 
hemolysis  of  human  corpuscles  by  cobra  venom.'*"  After  splenectomy 
there  is  an  increased  resistance  to  venom  hemolysis.^"'^ 

Eel  serum  is  remarkably  hemolytic,  so  much  so  that  a  quantity  of 
0.1  c.c.  per  kilogram  of  body  weight  will  kill  a  rabbit  or  guinea-pig 
in  three  minutes  when  injected  intravenously.  Heating  at  5-4°  C. 
for  fifteen  minutes  destroys  the  hemolytic  action,  and,  unlike  ordinary 
serum  hemolysins  the  addition  of  complement  does  not  restore  its  ac- 
tivity. Animals  can  be  immunized  against  this  serum.  Introduced 
into  the  stomach  in  ordinary  quantities  eel  serum  is  not  toxic.  It  can 
be  dried  and  redissolved  without  losing  its  activity,  but  acids  and 
alkalies  readily  destroy  it.  Mosso,  who  first  discovered  the  toxicity  of 
eel  serum,  called  the  unknown  active  principle  ichthyotoxin.  ]\Iany 
other  animals  produce  hemolytic  poisons  (e.  g.,  spiders,  bees)  which 
are  diseussetl  under  Zootoxins,  Chapter  vi. 

HEMOLYSIS  IN  DISEASE 

During  health  there  is  always  going  on  a  certain  amount  of  de- 
struction of  red  corpuscles  that  have  outlived  their  usefulness;  hence 
in  disease  we  may  have  to  deal  with  either  an  alteration  in  the  nor- 
mal processes  of  blood  destruction  or  the  introduction  of  entirely  new 
processes.  Although  the  place  and  manner  of  normal  red  corpuscle 
destruction  is  not  completely  known,  yet  it  seems  probable  that  there 
is  relatively  little  hemolysis  wathin  the  circulating  blood.  When  a 
red  corpuscle  becomes  damaged,  it  seems  to  become  more  susceptible 
to  phagocytosis,  and  it  is  then  picked  out  of  the  blood,  chiefly  by  the 
endothelial  cells  of  the  sinuses  of  the  liver,  spleen,  hemolymph  glands, 
and  bone-marrow.  Within  these  cells  it  apparently  undergoes  hemo- 
lysis. Eventually,  the  resulting  pigment  is  split  up  by  the  liver,  the 
non-ferruginous  portion  forming  the  bile-pigments,  while  the  iron 
seems  to  be  mostly  withheld  to  be  worked  over  into  new  hemoglobin.*"'' 

79  Jour.  Infect.  Dis.,  1909  (6),  688;  Stone  and  Schottstaedt,  Arch.  Int.  :Med., 
1912   (10),  8. 

80  See  articles  on  this  siibject  in  the  !Miinch.  med.  Woch.,  1909,  Vol.  .56. 
soaKolmer,  .Tonr.  Exp.  Med.,   1917    (25),   195. 

sobMuir  and  Dunn    (Jour.   Path,  and   Bact,   1915    (20),   41),  find  that  after 


230  CHEMISTRY    OF    THE    IMMUMTY    h'EACTIOXS 

(See  ' ' Pigineutation, "  Chap,  xvi.)  Wlioiievor  during  disease  red  cor- 
puscles are  more  rapidly  injured  than  they  are  under  normal  condi- 
tions, these  processes  of  normal  hemolysis  are  exaggerated  and  we  not 
only  find  the  phagoc.vtic  cells  of  tlie  spleen  and  glands  packed  with 
corpuscles,  but  endothelial  cells  elsewhere,  and  also  leucocytes,  take 
on  the  hemolytic  function.  At  the  same  time  there  results  an  exces- 
sive production  of  bile-pigment  from  the  destroyed  red  corpuscles, 
which  has  an  etiological  relation  to  the  so-called  "  hemato-hepatogen- 
ous"  jaundice.  If  hemolysis  is  very  excessive,  the  blood  pigment  ac- 
cumulates in  other  organs  than  the  liver  and  spleen.  According  to 
Pearce  *^  and  his  associates,  when  the  blood  contains  at  one  time  more 
than  0.06  gm.  of  free  hemoglobin  per  kilo  of  body  weight,  it  begins  to 
be  excreted  by  the  kidneys ;  smaller  amounts  are  cared  for  chiefly 
by  the  liver,  and  even  when  much  larger  amounts  of  hemoglobin  are 
present  in  the  blood  the  liver  takes  care  of  most  of  it,  only  a  rela- 
tively small  proportion,  17  to  36%,  being  excreted  in  the  urine. 
Hence  it  is  possible  to  have  hemolytic  jaundice  without  hemoglobin- 
uria. Part  of  the  pigment  is  converted  into  urobilin,  and  the  amount 
of  this  pigment  in  the  stool  is  an  index  of  the  amount  of  hemolysis.^^* 
In  persons  with  hemolytic  hemoglobinemia,  intravenous  injection  of 
hemoglobin  will  produce  hemoglobinuria  with  snuiller  dosage  than  in 
normal  persons,  who  require  at  least  17  c.c.  of  laked  corpuscles  to  pro- 
duce hemoglobinuria.^^'' 

It  is  possible  that  the  globin,  which  is  quite  toxic  when  f  ree,^-  may 
play  a  part  in  the  symptomatology  of  hemolytic  poisons.  The  stroma 
of  the  erythrocytes  also  seems  to  be  toxic. ^-^ 

The  resistance  of  erythrocytes  to  hemolytic  agents  varies  greatly 
in  disease  conditions  ^^  and  often  specifically, — i.  e.,  resistance  may  be 
increased  to  one  agent,  decreased  for  another,  and  normal  with  a 
third.  Attempts  have  been  made  to  use  this  resistance  as  a  diagnostic 
or  prognostic  index,  but  not  with  great  success  in  most  cases.  A])- 
parently  changes  in  the  plasma  lead  to  alterations  in  the  permea- 
bility of  the  corpuscles,  which  determines  their  behavior  with  hemo- 
lytic agents ;  also  changes  in  the  proportion  of  lipoids  and  hemo- 
globin may  modify  hemolysis.  As  an  example  of  this  condition  may 
be  cited  observations  on  hemolysis  by  cobra  venom,  the  corpuscles  hav- 
ing been  found  less  resistant  in  dementia  precox,  more  resistant  in 
carcinoma  and   sy])hilis.     Hutler®*   states   that   fragility   of   the   cor- 

acute  li(»ni()l\iic  niicTiiia  in  raliliils  llio  oxooss  iron  stored  in  tlic  oryans  has  liccii 
nearly  all  alisorbod  by  tlie  time  re<i'oneratioii  of  tlio  blood   is  coniplcU'. 

81  .lour.  Kxp.  !\Iod.,"ini2    (10),  several  articles. 

siaSco   llobeitsoii.   Areh.    Int.   IMed.,    101.')    (If.),    1072. 

«ib  Sellards  and  Minot,  Jour.  Mod.  Res.,  1010   (.34).  400. 

f<2  Reliittoiditdm  and  Woiohardt.  ISliincli.  mod.  Wooli.,   1012    (.")0).   lOSO. 

82aBarratt  and  Yorko,  Brit.  Mod.  .Tour.,  Jan.  .31.   1014. 

83  Review  by  Paltanf.  Krolil  and  :\lareliaiurs  Handb.  all-r.  rath(d..  l'.)12  (11 
( 1 )  1 ,  8.3. 

S'J  Quarterly  Jour.  Mod.,    1013    (0),   14."). 


II i:\lOLYSlH  J\  DISEA.Sl-J  231 

puscles  is  abnorinally  liigh  in  exophthalmic  f^oiter,  cancer,  syphilis, 
tabes,  anemia  and  malaria.  In  obstructive  jaundice  the  corpuscles 
show  an  increased  resistance  to  hemolysis  by  hypotonic  salt  solution, 
but  in  congenital  hemolytic  jaundice  the  resistance  is  decreased.*^* 
Using  saponin  hemolysis,  Bigland  ^*^  found  the  resistance  greatly  de- 
creased in  icterus,  although  the  serum  had  an  increased  protective 
action  because  of  antagonism  between  the  saponin  and  the  bile  salts; 
in  all  anemias  resistance  was  found  increased,  except  pernicious 
anemia,  which  showed  normal  or  slightly  subnormal  resistance ;  high 
temperature  decreases  resistance.  As  will  be  seen  from  the  few  ex- 
amples cited,  the  resistance  to  different  hemolytic  agents  may  vary 
with  the  same  corpuscles.**'' 

The  hemolysis  of  the  acute  febrile  diseases  is  readily  explained  by  the 
demonstrable  hemolytic  property  of  the  products  of  the  organisms 
that  cause  them,  such  as  streptocolysin,  staphylolysin,  etc.  Perhaps 
at  the  same  time  products  of  altered  metabolism  may  also  play  a 
part,  but  it  does  not  seem  probable  from  experimental  results  that 
the  thermic  condition  per  se  has  much  effect.  In  malaria,  although 
the  parasites  enter  and  destroy  the  corpuscles  in  which  they  live,  yet 
this  alone  does  not  account  for  all  the  blood  destruction  of  the  dis- 
ease, for  the  amount  of  anemia  is  quite  without  relation  to  the  num- 
ber of  parasites  to  be  found.  There  is  good  reason  to  believe  that  the 
Plasmodia  produce  hemolytic  substances  that  are  discharged  into  the 
serum.  In  the  primaiy  anemias  hemolysis  seems  to  be  the  essential 
process,  although  the  agents  involved  are  at  present  unknown.  Ab- 
sorption of  hemolytic  products  of  intestinal  putrefaction  or  infection 
has  always  come  in  for  much  suspicion,  without  ever  becoming  com- 
pletely established.  Here  also  the  hemolysis  seems  to  take  place  in 
the  endothelial  cells  rather  than  in  the  vessels.  In  such  a  disease  as 
pernicious  anemia  there  is  much  reason  to  assume  that  defective  or 
abnormal  hematogenesis  is  an  important  factor.  Probably  the 
anemia  of  nephritis  is  the  result  of  hemolytic  action  of  the  retained 
products  of  metabolism,  in  which  connection  the  hemolytic  properties 
of  ammonium  compounds  may  be  recalled.  In  some  diseases  asso- 
ciated with  anemia  it  has  been  found  that  the  blood-serum  of  the 
patient  is  distinctly  isohemolytic,  although  isoagglutination  seems  to 
be  more  frequent.  The  fluids  that  can  be  obtained  from  cancers  have 
been  found  to  be  hemolytic,  while  antihemolysin  has  been  found  in 
ascitic  and  pleural  effusions.  Autolytic  disintegration  of  liver,  and 
presumabl}'  other  tissues,  may  also  cause  the  presence  of  hemolytic 
substances    in    the    blood. ®^     Arseniuretted    hydrogen    may    produce 

84a  See  Richards  and  Johnson,  Jour.  Amor.  Med.  Assoc,   1913    (01).   15S0. 

84b  Quart.  .Tour.  Med..   1914    (7),  .370. 

84c  Bihliofrrapliy  by  Krasny.   Folia   Ilcmatol.,   1913    (16).   353. 

85  Maidorn,  Bioehem.  Zeit.,  1912  (4.5),  328.  Hemolytic  lipoids  are  believt-d  by 
some  to  )ie  liberated  from  injured  tissues  (see  Kirsche.  Bioehem.  Zeit.,  1913  (ol). 
169),  but  McPhedran   (Jour.  Exp.  Med.,  1913    (18),  527)    could  find  no  evidence 


232  CHEMISTRY    OF    THE    liJilUMTY    h'EACTIOXS 

hemolysis  in  some  such  way,  since  it  causes  no  hemolysis  in  the  test 
tube  (Ileffter).  The  very  great  hemolytic  action  of  soaps  and  free 
fatty  acids,  which  varies  directly  with  the  number  of  unsaturated 
carbon  atoms  they  contain  (Moore  **"),  makes  it  possible  that  these  sub- 
stances play  a  part  in  the  hemolysis  of  disease,  especially  since  the 
fatt}'  acids  of  the  liver  are  characterized  by  their  high  content  of  free 
fatty  acids.  Bile  is  strongly  hemolytic,  and  in  icterus  this  is  an  im- 
portant consideration. 

In  many  forms  of  poisoning  hemolysis  is  a  prominent  feature ;  in 
some  it  seems  to  be  the  chief  effect  of  the  poison,  e.  g.,  potassium 
chlorate  and  arseniuretted  hydrogen.  In  severe  extensive  burns  there 
may  occur  hemolysis,  and  hemoglobinuria  may  also  result.  The  hemo- 
globinuria of  "blackwater  fever"  has  been  the  cause  of  much  discus- 
sion as  to  whether  the  malarial  parasite  or  the  quinine  is  the  cause, 
with  a  divided  opinion  resulting,  although,  undoubtedl}',  cases  do  occur 
in  malaria  without  administration  of  quinine.  The  studies  of  Brem  *■ 
indicate  that  the  hemolysis  is  produced  by  a  hemolysin  coming  from 
the  Plasmodium,  and  that  the  quinine  influences  the  condition  by  pre- 
venting the  action  of  an  antihemolytic  substance  present  in  the  blood. 

After  removal  of  the  spleen,  hemolysis  by  the  hemolymph  glands  ex- 
ceeds that  of  the  primitive  spleen,  causing  an  excessive  destruction  of 
red  corpuscles  (Warthin  ***).  This  suggests  that  the  spleen  may  nor- 
mally dispose  of  some  hemolytic  agent  which  acts  either  by  stimu- 
lating phagocytosis  or  by  so  altering  the  red  cells  that  they  are  par- 
ticularly susceptible  to  phagocytosis.  This  idea  is  not  substantiated 
by  the  work  of  Pearce,^**  who  found  the  anemia  of  splenectomy  ac- 
companied by  an  increased  resistance  of  the  corpuscles  to  hemolysis, 
and  no  hemolytic  agent  was  present  in  the  blood.  There  also  occurs 
the  group  of  anemias  associated  with  great  enlargement  of  the  spleen, 
and  in  which  removal  of  the  spleen  ma}'  result  in  a  return  to  normal 
blood  conditions;  a  fact  suggesting,  among  other  possibilities,  that 
there  may  be  poisons  which  stimulate  directly  the  hemolj'tic  action  of 
the  spleen  independent  of  the  natural  stimulation  of  splenic  hemolysis, 
which  comes  from  the  presence  in  the  splenic  blood  of  injured  red 
corpuscles.*'"^ 

Paroxysmal  Hemoglobinuria.'"' — This  condition  seems  to  depend 
upon  the  presence  in  the  serum  of  a  hemolytic  amboceptor  (an  auto- 

that  any  particularly  licmolytic  fatty  afid,  more  active  (lian  oleic  acid,  can  be 
isolated  from  either  normal  or  diseased  tissues. 

s«  Brit.  Med.  Jour.,  1909  (ii),  0S4;  see  also  Lamar.  Jour.  Kxper.  Med.,  1911 
(13),  380. 

STArcli.   Int.  Med.,    1912    (9),    129. 

88  Jour.  Med.  Researcii,   1902    (7),  4.'{.'). 

89  Pearce  ct  at.,  Jour.  Kxp.  Med.,  1912,  \'ol.  IG.  Sec  also  Roccavilla,  Arch.  Mtnl. 
Exp.,  191.5    (26),  508. 

*<»aSee  Ranti,  Semain  Med.,  1913    (33),  3i:?. 

'•'O  T.andsteiner,  IIandl)Uch  d.  Riochem.,  \'ol.  2  ( 1 ) .  ]).  492;  Me\(M-  and  iMniiiericli. 
Deut.  Arch.  klin.  Med.,  1909   (96),  287. 


HEMOLYSIS  I\   IHSEASE  233 

hemolysin),  which  Avill  eunibine  with  tlic  corpuscles  of  the  same  indi- 
vi(hial  and  sensitize  them  for  his  own  complement  (Donath  and  Land- 
steiner,  Eason).  This  antohemolysin  can  react  w^th  the  corpuscles 
only  at  low  temperature,  such  as  nuiy  be  furnished  in  the  peripheral 
vessels  by  exposure  to  cold,  and  the  complement  unites  when  the 
temperature  of  these  corpuscles  again  reaches  37°  in  other  parts- 
of  the  body.  In  susceptible  persons  attacks  of  hemoglobinuria  may  be 
brought  on  merely  by  holding  the  hands  in  cold  water,  and  their  blood 
senim  will  sensitize  to  hemolysis  human  corpuscles  (even  of  normal 
individuals)  in  vitro  at  low  temperatures. ""''  Certain  infections,  es- 
pecially syphilis,"^  predispose  to  paroxysmal  hemoglobinuria.  Not 
only  the  hemolytic  amboceptors,  but  also  an  auto-opsonin  is  present 
(Eason)  and  the  resistance  of  the  red  corpuscles  is  decreased  to  various 
harmful  agencies,  including  CO,  and  otiier  weak  acicls.^-  The  cor- 
puscles of  three  cases  studied  by  jMoss  '^^  showed  an  increased  resistance 
to  hypotonic  NaCl  solutions.  Just  before  the  rigor,  hemolysins  may 
be  found  in  the  blood,  disappearing  after  the  hemoglobinuria  (Rob- 
erts)."^'"' In  a  case  studied  by  Dennie  and  Robertson,^^"  hematuria 
resulted  from  destruction  of  only  6.3  c.c.  of  the  patient's  blood,  and 
90  per  cent,  of  the  liberated  hemoglobin  was  excreted  within  two  hours. 
Pathological  Anatomy. — The  lesions  produced  in  the  organs  of 
animals  injected  with  hemolytic  agents  are  usually  pronounced  and 
quite  characteristic.  There  is  often  a  subcutaneous  edema,  which 
is  usually  blood-stained,  and  similar  'fluid  may  be  present  in  the 
serous  cavities.  The  fat  is  yellowish,  and  the  muscles  are  darker  in 
color  than  is  normal.  The  spleen  is  usually  much  swollen,  soft,  fria- 
ble, and  very  dark  in  color.  The  liver  is  usually  sw^ollen  and  mottled 
with  red  areas  in  a  yellow  background.  The  renal  cortex  is  dark  in 
color,  even  chocolate-colored,  and  the  pyramids  are  comparatively 
light;  hemoglobin  is  freqiiently  present  in  the  urine.  In  the  lungs 
are  often  found  hemorrhages  or  areas  resembling  small  infarcts.  The 
blood  may  be  thin  and  even  distinctly  transparent.  Microscopically 
the  red  corpuscles  are  found  in  all  conditions  of  degeneration,  and 
often  fused  together.  In  the  liver,  besides  patches  of  congestion, 
fatty  changes  are  present  if  the  animal  lives  long  enough.  Large 
phagocytic  cells  packed  with  red  cori3uscles  are  abundant  in  the  spleen 
and  lymph-glands,  as  well  as  diffuse  accumulations  of  the  blood.-cells, 
which  are  often  fused;  and  much  pigment  is  also  present,  both  free 
and  in  the  cells.  Pigment  also  accumulates  in  the  renal  epithelium, 
which   often    shows   much    disintegration ;    congestion    is    prominent, 

90a  Widal    looks    upon    paroxysmal    hemoglobinuria    as    an     autoanaplivlaxis- 
{Semain  Med.,  1913   (.3.3),  613). 

91  Matsuo,  Arch.  f.  klin.  Med.,  1912    (107)    335. 

92  Berghausen,   Arch.   Int.   Med.,    1912    (9),    137. 

93  Folia  Serologica,  1911    (7).  1117. 
93a  Brit.  Med.  Jour.,   1915    (2),  398. 
93b  Arch.  Int.  Med.,  1915    (16),  205. 


234  CHEMISTRY    OF    THE    JMMUyiTY    JiEACTIOXS 

and  hemorrhages  iuto  both  interstitial  tissue  and  giomerules  are  fre- 
quent. Some  of  the  lesions  are  due  to  the  hemolysis,  and  some  to  the 
associated  agglutination  of  corpuscles,  which  form  hyaline  thrombi. 
Pearce  '•'*  has  found  that  agglutinative  serum  when  injected  into  dogs 
causes  widespread  necrosis  in  the  liver,  which  is  followed  by  prolifera- 
tion of  connective  tissue  and  the  production  of  changes  resembling 
cirrhosis.  There  is  a  marked  decrease  in  the  gl^'cogen  content  of  the 
liver,  and  of  its  lipolytic  activity   (Andrea)."^ 

COMPLEMENT  FIXATION  "■  AND  WASSERMANN  REACTIONS" 

The  original  principle  involved  in  these  reactions  was  first  demon- 
strated by  Bordet  and  Gengou,  and  is  essentially  as  follows :  If  a 
specific  antigen  and  amboceptor  unite  in  the  presence  of  complement, 
the  complement  is  then  united  to  the  amboceptor-antigen  compound  to 
complete  the  reaction.  AVhen  sufficient  amounts  of  amboceptor  and 
antigen  are  present  the  entire  quantity  of  available  complement  may 
be  thus  fixt,  and,  consequently,  the  mixture  contains  no  more  comple- 
ment for  further  reactions.  As  complement  does  not  ordinarily  unite 
v/ith  amboceptors  except  when  the  amboceptors  are  united  with  their 
specific  antigens,  the  fact  that  in  a  given  system  of  complement 
-(-  amboceptor  -|-  antigen  there  is  no  free  complement,  is  evidence 
of  a  reaction  between  amboceptor  and  antigen ;  in  consequence  of 
which  this  reaction  can  be  used  to  determine  the  presence  of  a  specific 
amboceptor  in  a  serum,  by  using  the  corresponding  antigen ;  or  con- 
versely, with  a  serum  containing  a  known  amboceptor  we  can  detect 
the  presence  in  a  solution  of  the  specific  antigen.  The  indicator  of 
the  presence  or  absence  of  complement  which  is  in  universal  use,  is 
the  ability  of  the  mixture  to  hemolyze  erythrocytes  in  the  presence 
of  the  specific  hemolytic  amboceptor.  Thus,  if  typhoid  bacilli  and  a 
typhoid  antiserum  which  contains  both  complement  and  specific  am- 
boceptor, are  mixed  in  proper  proportions  and  incubated  for  a  short 
time,  the  complement  will  he  bound  to  the  bacilli.  If  we  then  add  this 
mixture  to  sheep  corpuscles  which  have  been  acted  ujion  by  an  anti- 
sheei)-corpuscle  serum,  from  which  the  complement  had  been  previously 

M.Toiir.  Exp.  Med.,  1906   (8),  64;  Jour.  :\r<'d.  l^seardi.  IIMKI   (14).  .-)41. 

95  Arch,  internat.  pharniaeodyn.,   100.5    (14),   177. 

96  Tlie  roaction  of  "complomont  fixation"  must  not  hv  confused  witli  tlio  en- 
tirolv  distinct  reaction  of  "coniiilemcnt  deviation,"  a  mistake  very  lil<ely  to 
liappen  because  of  the  careless  but  erroneous  use  by  some  writers  of  tlie  latter 
icnii  ill  describing  tlie  first-named  reaction.  Complement  deviation  (or  Xeisser- 
\\'cclisl)t"r^f  j)beii()iiienon)  is  juoduced  wlien  an  excess  of  aniboccplors  is  present 
tofXetlier  witli  antijren  and  a  limilcd  amount  of  coui]d('intMi(.  wliicli  results  in 
absence  of  complement  activity.  The  inccliaiiism  of  tliis  reaction  has  not  liccn 
satisfactorily  explained. 

9"  Literature  <riven  by  ^Feier,  .Tahresber.  d.  Tmmunitiitsforscli.,  1909  (4).  58; 
Sacbs  and  Altmann,  Kolle  and  Wasscrmann's  llandbucli.  Krfriin/unjjjsbd.,  2,  1909, 
ji.  47(>:  No^iiclii.  "Sciiiin  Diagnosis  of  Svpliilis  and  l.uclin  Head  ion."  I'liila- 
dclpliia,   1912. 


<'()Mri.i:\n:\T  fixation  and  wassermann  ke action       235 

reiuuved  by  licatiiiy,  no  liemolysis  will  occur,  i'or  we  have  added  no 
free  complement.  But  if  our  original  mixture  had  contained  dysen- 
tery hacilli  instead  of  tyi)lioid  bacilli  the  complement  would  not  have 
been  fixed,  and  the  addition  of  this  mixture,  containing  free  comple- 
ment, to  the  sensitized  sheej)  eoi'puseles  would  cause  prompt  hemol- 
ysis. 

This  reaction  was  at  first  used  for  the  detection  of  antibodies  in 
sera,''*  and  for  the  identification  of  bacteria,  and  was  found  to  be  ex- 
(iuisitely  delicate,  detecting  most  minute  amounts  of  antigens  with 
the  sharpest  specificity  limits  of  any  of  the  immunity  reactions.  On 
account  of  the  delicacy  of  this  reaction  it  can  be  used  to  determine 
the  presence  in  tissues  of  specific  organisms  which  cannot  be  culti- 
vated ;  thus,  it  has  been  possible  to  demonstrate  the  existence  of  a 
specific  scarlatinal  virus ""  in  the  tissues  during  this  disease,  although 
the  actual  organism  cannot  be  isolated.  This  fact  led  Wasser-inann 
to  use  extracts  of  the  livers  of  congenital  syphilitic  fetuses,  which 
contain  great  quantities  of  spirochetes,  as  an  antigen  for  complement 
fixation  reactions,  whereby  it  should  be  possible  to  determine  in  a 
given  serum  the  presence  of  specific  amboceptors  for  the  virus  of 
syphilis,  such  amboceptors  being  present  in  persons  infected  with 
syphilis  as  a  result  of  the  reaction  to  the  infection.  As  originally  in- 
troduced, then,  the  Wassermann  reaction  was  supposed  to  be  simply 
a  specific  reaction  between  syphilitic  antigen,  specific  syphilitic  am- 
boceptors, and  non-specific  complement.  It  was  soon  learned,  how- 
ever, that  the  reaction  as  it  occurred  in  syphilis  was  decidedly  dif- 
ferent from  the  original  complement  fixation  reaction  of  Bordet  and 
Gengou,  for  it  was  found  possible  to  substitute  in  the  reaction  for 
extracts  of  tissues  containing  syphilitic  virus  (spirochetes),  the  most 
varied  sorts  of  tissue  extracts,  coming  from  tissues  certainly  free 
from  spirochetes  (e.  g.,  ox  heart).  Noguchi  and  Bronfenbrenner  ^ 
summarize  the  i)resent  state  of  the  matter  in  these  words:  "We 
know  merely  this :  that  complement  in  the  presence  of  syphilitic  anti- 
gen may  be  rendered  inactive  by  one  or  more  substances  in  the  body 
fluids  of  a  syphilitic  or  parasyphilitic  patient." 

Extended  investigation  of  these  non-specific  antigens  whicli  give 
specific  complement  fixation  with  syphilitic  sera,  has  shown  them  to 
be  related  to  the  lipoids,  especiallj"  the  lecithins,  as  indicated  by  the 
fact  that  the  most  efficient  antigens  contain  the  aceton-insoluble  frac- 
tion  of  the  tissue  lipoids.     The  antigenic  value  of  this   fraction   of 

3S  Accordinfi  to  Gay  (I^niv.  of  Calif.  Publ.,  rathol.,  mil  (2),  ],  full  dispus- 
s-on)  complement  fixation  is  jirodueod  by  an  antioen-antihody  complex'  distinct 
from  precipitino<;en-j)reci[)itin,  l)ut  Dean  (Zeit.  f.  Tmnuinitiit.,  1012  (1.31,  S4)  be- 
lieves that  they  represent  two  phases  or  stages  of  tlie  same  reaction.  Thiele  and 
Embleton  (Zeit.  Tmmnnitiit.,  I!tl3  (10),  4.10)  consider  tliat  in  sypliilis  it  is  not 
a  specific  antibody,  but  an  anti-complementary  sul)stance  wliicli  arises  from  the 
disintefjrating  tissues. 

on  Koessler  and  Koessler,  .Tour.  Tnfec.  Dis.,   1012    (!)).  .30(1. 

iJour.  E.\-p.  Med.,   1911    (13),  43. 


236  CHEMISTRY    OF    THE    IMMUNITY    REACTIONS 

different  liver  extracts  varies  nearly  directly  with  its  power  to  com- 
bine with  iodin  -  (Noguchi  and  Bronfenbrenner) ,  which  indicates 
that  the  unsaturated  fatty  acids  are  important  in  the  reaction.^ 
Lecitliins  from  different  sources  vary  in  efficiency,  heart  lecithin  being 
jnore  active  than  liver  lecithin,  brain  and  egg  yolk  lecithin  following. 
Addition  of  cholesterol  to  the  lecithin  solutions  increases  greatly  their 
activity.*  An  acetone-precipitated  "antigen"  of  this  class  is  not  a 
true  antigen,  however,  for  fixation  antibodies  are  not  developed  in 
aninuils  injected  witli  such  a  lipoid  which  has  been  shown  to  be  en- 
tirely efficient  in  the  Wassermann  reaction.'^ 

As  for  the  substance  in  the  syphilitic  serum  which  participates  in 
the  Wassermann  reaction,  it  w-ould  seem  to  be  related  to  the  globulins, 
which  are  decidedly  increased  in  the  blood  and  spinal  fluid  '^  of 
sypliilitics,'''^  especially  the  euglobulin.'  P.  Schmidt  ^  ascribes  the 
reaction  to  the  physico-chemical  properties  of  the  globulins  of  the 
syphilitic  serum,  which,  he  believes,  possess  a  greater  affinity  for  the 
colloids  of  the  antigen  than  normal  globulins;  this  affinity  is  held  in 
check  in  normal  serum  b}'  the  albumins  of  the  serum,  which  are  rela- 
tively or  absolutely  decreased.  That  physico-chemical  factors  do  play 
a  part  is  evidenced  by  the  common  observation  that  the  turbidity  of 
the  antigen  suspension  is  closely  related  to  its  efficiency,  clear  solu- 
tions being  less  active.  Slight  changes  in  H-ion  concentration  will 
change  a  reaction  from  negative  to  positive,  or  reverse;  and  neutral 
salts  can  change  a  negative  to  a  positive  reaction,  but  not  the  reverse 
(Gumming).'*'^  The  lipoids  in  syphilitic  sera  are  said  by  Peritz  ^  to  be 
increased,  but  the  lipoid  content  and  the  antibody  titer  do  not  show 
any  constant  relation  (Bauer  and  Skutezky ).■'-''  The  cholesterol  con- 
tent of  syphilitic  blood  shows  no  evidence  of  a  quantitative  relation  to 
the  Wassermann  reaction.'*''     Friedemann  ^°  believes  that  a  globulin- 

2  Not  corroborated  by  Browning,  Cniioksliank  and  Gilmour. 

3  An  interesting  obsorvation  made  by  Xognclii  and  Bronfenbrenner.  is  that  ex- 
tracts frona  fatty  livers  arc  almost  devoid  of  antigenic  properties;  but  So  (Cent.  f. 
Bakt.,  1912  (63),  438)  found  that  the  extract  from  fatty  hearts  of  guinea-pigs 
was  more  active  than  from  normal  hearts. 

4  Browning  et  al.,  Zeit.  Immunitiit.,  1012  (14).  284;  Jour.  Pathol,  and  Bact., 
1911  (16),  135  and  225.  Klein  and  Fraenkel  believe  the  "antigen"  of  ox  heart 
extracts  to  be  a  comltination  of  lecithin  with  cliolosterol  and  small  amounts  of 
a  soap-like  substance  similar  to  jecorin    (IMiincli.  mcd.  \Voch.,  1914    (61),  651). 

5  Fitzgerald  and  LeaOics.  Tniv.  of  Calif.  I'ul)!.,   Path.,   1912    (2),  39. 
ePfein'er,  Kol)cr  aiid  Field,  Broc.  Soc.  Exp.  Biol.,   1915    (12),  153. 
oaSee  Kowe,  Arch.  Int.  ^Nled.,  1916   (18),  455. 

TMiiller  and  Hough,  Wien.  klin.  Woch.,  1911    (24),  167. 

8  Zeit.  f.  Hvg.,  1911  (69),  513.  See  also  Hirschfeld  and  Klinger,  Zeit.  Immuni- 
tUt.,  1914   (2i),  40. 

8a  Jour.  Infect.  Dis.,  1916    (18),  151. 

0  Zeit.  exp.  Path.,   1910    (8),  255. 

'JaWicn.  klin.  Wodi.,  1913    (26),  830. 

«b  Weston,  Jour.  Med.  Pes.,  1914  (30),  377;  Stein,  Zeit.  exp.  Med.,  1914  (3), 
309. 

10  Zeit.  f.  Ilvg.,  1910   (67),  279. 


COMPLEMEXT  FIXATIOS  AM)    W  ASSEltM WX   REACT/OX         237 

soap  conipouiid  is  the  active  substance  in  sypliilitic  sera.  ^Mcintosh  " 
says  that  the  active  component  differs  from  typical  antibodies  in  not 
passing  through  collodion  or  porcelain  filters,  and  there  are  many  who 
hold  that  the  reacting  substance  is  a  product  of  tissue  disintegration. 
AVassermann  ^^'"^  has  found  evidence  that  the  antibody  is  derived  from 
the  lymphocytes,  at  least  in  the  spinal  fluid  of  syphilitics. 

Whether  true  antibodies  are  concerned  in  the  Wassennann  reaction 
is  a  question.  In  favor  of  this  view  is  the  fact  that  the  serum  of 
rabbits  immunized  with  congenital  syphilis  livers  contains  an  anti- 
body giving  the  Wassennann  reaction,  exactly  like  the  serum  of 
syphilitics.'-  On  the  other  hand,  the  actual  substance  of  pure  cultures 
•of  spirochetes  does  not  ordinarily  act  as  antigen  with  syphilitic  sera 
in  the  Wassennann  reaction  (Noguchi).  It  is  highly  probable  that 
when  syphilitic  liver  extracts  are  used  as  antigen  in  the  Wassermann 
reaction,  we  have  a  true  Bordet-Gengou  reaction  of  complement  fixa- 
tion with  the  syphilitic  substance  present  in  this  extract,  in  addition 
to  the  reaction  which  is  accomplished  by  the  lipoids.  Whether  the 
complement  is  destroyed  by  enzymes,^^  or  is  inhibited  by  anti-comple- 
ment present  in  syphilitic  serum,  or  is  destroyed  by  some  toxic  sub- 
stance in  the  serum  "  are  matters  still  under  discussion.  A  favorite 
interpretation  of  the  Wassermann  reaction,  which  seems  to  harmonize 
with  the  known  facts,  is  that  there  is  a  precipitation  of  serum  globu- 
lin by  the  lipoidal  colloids  of  the  antigen,  and  adsorption  of  the  com- 
plement by  this  precipitate. 

Klausner's  Serum  Reaction. — When  distilled  water  is  added  in  cer- 
tain proportions  to  fresh  serum,  a  distinct  flocculent  precipitate 
separates  out  in  a  few  hours,  and  this  property-  is  much  more  marked 
in  syphilitic  than  in  normal  sera.  While  not  specific  for  syphilis,  this 
reaction  is  almost  invariably  present  in  certain  stages  of  syphilis. 
This  property  is  not  due  to  the  excess  of  globulin  present  in  sj^phi- 
litic  sera,  according  to  the  later  studies  of  Klausner,^^  who  be- 
lieves that  the  high  lipoid  content  of  syphilitic  serum  is  responsi- 
ble.'-^ 

Porges-Hermann-Perutz  Reaction. — If  equal  parts  of  a  2%  solution 
of  sodium  glycocholate  and  an  alcoholic  cholesterol  suspension  (0.4%) 
<ire  added  to  inactivated  serum  from  syphilitic  patients,  a  precipitate 

11  Zeit.  Immunitiit.,  1910   (5),  76. 

iia  Wassermann  and  Lange,  Berl.  klin.  Woch..  1014   (51),  .527. 

12  Citron  and  Munk,  Deut.  med.  Woch.,  1910  (36),  1560:  Eiken.  Zeit.  Imninni- 
tat.,  1915   (24),  188. 

13  Manwarinjr,  Zeit.  f.  Immunitiit..  1909    (.3),  309. 
1*  Kiss,  ihid.,   1910    (4),  703. 

15  Biochem.  Zeit.,  1912    (47),  36. 

15a  Tliat  the  serum  of  sypliilitics  is  much  altered  chemieally  is  shown  by  all 
these  reactions.  Bruck  has  also  found  that  the  precipitate  produced  by  adding 
strong  nitric  acid  to  syphilitic  serum  is  characteristically  insoluble  and  gelatinous 
(Miinch.  med.  Woch.,*  1917    (64),  25). 


238  CHEMISTRY    OF    TllTJ    IMMUMTY    REACTIOXS 

forms,  while  with  normal  serum  there  occurs  no  precipitate.^*^     Little 
is  known  concerning  the  nature  of  this  reaction. 

Coagulation  Reaction. — This  was  described  by  Hirschfeld  and 
Kling-er,^'^''  and  depends  on  the  fact  that  tissue  extracts  digested  with 
syphilitic  serum  lose  tlieir  ability  to  coagulate  blood.  The  effect  is 
believed  to  depend  on  adsorption  of  the  lipoids  of  the  tissue  extract  by 
serum  constituents,  and  henee  is  fundamentally  similar  to  the  Wasser- 
mann  reaction. 

CYTOLYSIS  IN  GENERAL  '' 

Not  the  same  degree  of  success  has  been  obtained  in  immunizing 
against  otlier  tissue  elements  as  with  the  erythrocytes.  Immune 
serum  can  readily  be  obtained  against  cells  that  can  be  secured  quite 
free  from  other  cells,  such  as  spermatozoa,  ciliated  epithelium,  and 
leucocytes,  but  even  then  the  immunity  is  not  specific.  ]\luch  less  is 
it  specific  when  ground-up  organs  are  used  for  innnunizing,  as  is  the 
case  in  the  experimental  production  of  iieplirolysins,  hepatolysins, 
etc.,  for  at  the  same  time  antibodies  are  secured  for  not  only  the 
typical  parenchyma  cells,  but  also  for  endothelium,  stroma  cells,  red 
and  white  corpuscles,  and  blood  plasma.  As  a  consequence,  the  early 
expectations  that  by  this  process  of  immunization  against  specific  cells 
great  progress  could  be  made  in  our  knowledge  of  physiology,  by 
selectively  throwing  out  of  function  an  organ  through  the  simple 
process  of  injecting  an  antiserum,  have  been  disappointed.  Equally 
little  progress  has  been  made  in  the  treatment  of  malignant  growths 
by  the  same  method.  The  immune  serums  usually  obtained  do,  to 
a  certain  extent,  injure  the  specific  organ,  but  tliey  also  usually 
injure  other  organs  nearly  as  much  or  perhaps  more ;  furthermore  they 
generally  contain  hemolytic  toxins,  even  if  the  tissues  used  in  im- 
munizing are  free  from  blood,  and,  as  we  have  seen,  hemolytic  poisons 
may  cause  serious  tissue  destruction.^^ 

Beebe  ^^  claims  to  have  secured  serums  by  immunizing  with  tissue 
iiucleoproteins,  that  were  altogether  specifically  toxic  for  the  type  of 
cells  from  which  the  nucleoproteins  were  obtained;  e.  g.,  immunizing 
with  liver  nucleoproteins  .yielded  serum  destroying  liver  cells  and 
no  others.  Other  observers  have  failed  to  corroborate  this  work.-" 
According  to  Zinsser  -^  the  cytolytic  antibodies  may  be  quite  distinct 

16  See  Gammeltoft,  Dent.  mod.  Wooh.,  1912  (38),  10.34;  Ellermaim,  (7)/^/..  l!)i;i 
(39),  210. 

loaDput.  nied.  Wooh.,  1014    (40),  1007. 

1"  Literature  is  given  liv  Fieisclimann  and  Davidsolin,  Folia  Serologiea.  lOOS  (1), 
173;  Landsteiner,  Handinicli  d.  liioeliem.,  1000  (II  (1)  ),  542;  Riteliie,  Jour. 
Pathol,  and  Haet.,  1008    (12),  140. 

18  See  Sata,  Zieplor's  Beilr.,   1906    (39),   1. 

10  Jour.  Kx]).  Med.,  190ri    (7),  733. 

20  Poarco  and  Jaekson,  Jour.  Infeet.  Dis.,  lOOfi  (3),  742.  See  also  review  l)v 
Wells,  Zeit.  f.  Imnuinitiit..  1913    (10),  500. 

21  Hioeheni.  Zeit.,  lOKi   (77).  120. 


CYToLYsis  /\  (!i:si:iiM.  239 

from  tlie  albumciiolyl  ic  aiulxiceptors  wliicli  arc  dcvt'luped  afrainst  un- 
formed protein  antigens. 

In  view  of  the  present  uncertain  state  of  the  subject,  and  the  very 
questionable  value  of  much  of  the  work  so  far  done,  the  consideration 
of  the  various  cytolysins  or  cytotoxins  may  be  dismissed  by  briefl\- 
referrinp:  to  a  few  of  the  most  important  results. 

Leucocytolytic  Serum.-- — Tliis  may  be  obtained  either  by  immuniz- 
ing with  leucocytes  obtained  from  e.xudates  or  from  the  blood,  or  by 
using  emulsions  of  lymph-glands.  Specific  leucocytolytic  serum  ag- 
glutinates leucocytes  and  produces  observable  morphologic  changes, 
in  the  way  of  solution  of  the  cytoplasm  and  cessation  of  ameboid 
movements ;  but  it  may  also  react  with  the  fixed  tissue  cells  of  the  same 
animal.-^  Of  the  leucocytes,  the  large  granular  cells  seem  most  af- 
fected and  the  lymphocytes  least.  "When  injected  into  the  peritoneal 
cavity  such  serum  causes  an  apparent  initial  leucopenia.  and  later  a 
decided  leucocytosis  in  the  peritoneal  fluid.  Corresponding  with  this, 
if  bacteria  are  injected  at  the  same  time  as  the  serum,  resistance  is 
found  decreased,  but  later  it  is  much  increased.  Such  serum  also 
contains  anticomplement,  according  to  AVassermann,  indicating  that 
the  injected  leucocytes  contain  complement.  Leucocytotoxin  obtained 
by  immunizing  against  lymphatic  tissue  is  very  thermolabile,  being 
destroyed  by  55°  C.  for  thirty  minutes,  and  the  serum  can  be  only 
partially  reactivated  by  the  use  of  fresh  serum.  Bacterial  filtrates 
may  also  contain  "leucocidins"  analogous  to  hemolysins.  Normal 
foreign  sera  are  more  or  less  toxic  to  leucocytes,  which  can  be  shown 
by  the  reduced  capacity  of  the  leucocytes  for  phagocytosis.^^^ 

Antiplatelet  Serum. — Several  experimenters  have  produced  antisera 
for  platelets.  Lee  and  Robertson  ~^^  obtained  a  specific  lytic  and  ag- 
glutinative action,  requiring  complement  for  its  accomplishment.  In- 
jected into  animals  this  antiplatelet  serum  caused  a  condition  resem- 
bling exactly  purpura  liemorrliagica  in  man. 

Endotheliolytic  Serum. — Every  attempt  at  immunizing  an  animal 
with  any  sort  of  fixed  tissue  must  of  necessity  involve  the  injection 
of  endothelial  cells  as  well  as  the  cells  specific  to  the  tissue  studied. 
Therefore,  it  is  possible  that  cytotoxic  serum  so  obtained  will  contain 
endothelial  toxins,  and  so  complicate  any  results  of  intra  rt'tam  ex- 
periments. There  is  every  reason  to  believe  that  endotheliolytic  sub- 
stances are  produced  in  this  way.  Ricketts  found  that  serum  of  ani- 
mals immunized  against  lymph-glands  was  toxic  to  endothelial  cells, 
which  was  indicated  by  hemorrhages  at  the  point  of  injection,  and 

22  Literature,  see  Flexner,  Univ.  of  Penn.  "Nred.  Bull.,  1002    (15),  2S7 :  Eicketts, 
Trans.  Chicatjo  Path.  Soc.  1902   (5),  178:  Christian.  Deut.  Arch.  klin.  ^Med.,  1904 
(80),  333;  Leschke,  Zeit.  Immunitiit.,  1913    (16),  627;   Eeeser,  Folia  Mikrobiol., 
1914.  H.  3. 
23Spiit,  Zeit.  Tmmunitat..  1914   (21),  565. 
-    23aLohner,  Arch.  ges.  Phvsiol.,  1915   (162),  129. 
23b  Jour.  Med.  Res.,  1916  "(33),  323. 


240  CHEMISTRY    OF    THE    JMMUMTV    BE  ACTIONS 

marked  desquamation  of  endothelium  when  the  injection  was  made 
into  a  serous  cavity.  In  snake-venom  poisoning  the  extensive  hemor- 
rhages are  also  due  to  an  endotheliolytie  principle,  called  by  Flexner 
hemon^hagin. 

Lymphatolytic  Serum. — This  serum  has  been  studied  by  Ricketts 
and  by  Flexner,  who  immunized  animals  with  lymph-glands.  As 
might  be  expected  from  the  structure  of  the  injected  glands,  the  re- 
sulting serum  contained  endotheliotoxin,  leucocytotoxin,  hemolysin, 
hemagglutinin,  leucocyto-agglutinin,  and  precipitins.  When  injected 
into  animals,  this  serum  has  a  marked  effect  upon  the  spleen  and 
lymph-glands,  producing  great  enlargement  and  congestion  of  these 
structures.  The  bone-marrow  is  also  somewhat  affected,  and  when 
marrow  is  used  in  immunizing,  the  mj/eloto.ric  serum  produces  marked 
proliferative  changes  in  the  lymph-glands  as  well  as  in  the  marrow. 

Nephrolytic  Serum. — It  has  been  claimed  that  if  a  kidney  is  de- 
stroyed by  ligating  its  vessels  or  ureter,  the  remaining  kidney  de- 
velops serious  degenerative  changes,  which  are  not  present  if  one  kid- 
ney is  entirely  removed.  This  has  been  attributed  to  Ihe  development 
of  nephrotoxic  substances  produced  in  reaction  to  the  absorption  of 
the  injured  renal  tissue  that  has  been  left  in  the  body.  Other  methods 
of  renal  injury  have  been  thought  to  produce  similar  effects,  and 
serum  of  animals  with  kidney  disease  was  said  to  injure  the  kidneys 
of  normal  animals.  Upon  this  basis  it  has  been  thought  possible  to 
explain  the  progressive  nature  of  the  chronic  nephritides  as  the  result 
of  nephrotoxins  produced  through  the  absorption  of  the  injured  cells, 
wiiich  nephrotoxins  injure  still  other  renal  cells.-*  Such  a  process, 
however,  involves  the  production  of  cell  toxins  in  an  animal  that  are 
toxic  for  its  own  cells,  that  is,  autocytotoxins ;  and  as  it  has  so  far 
been  practically  impossible  to  produce  autolysins  of  other  sorts,  it  is 
not  altogether  probable  that  the  kidney  is  an  exception.  Further- 
more, Pearce  -'  was  unable  to  produce  isonephrotoxins,  and  could  not 
corroborate  the  statements  as  to  the  changes  said  to  have  been  found 
in  the  remaining  kidney  after  ligating  the  vessels  of  its  mate.  He 
did  obtain  an  active  heteronephrolysin,  but  also  found  that  immuni- 
zation with  liver  produced  nearly  as  actively  nephrolytic  serum  as 
did  innnunization  with  kidney. 

Neurolytic  Serum. — Even  as  highly  s])ecialized  cells  as  those  of 
the  nervous  tissue  seem  to  produce  a  reaction  with  the  formation  of 
iimnnne  bodies.  Perhaps  the  most  positive  results  are  those  of 
Ricketts  and  Rothstein,-''  who  found  that  serum  of  rabbits  immunized 
against  the  brains  or  cords  of  guinea-pigs  was  highly  toxic  when  in- 
jected into  the  vessels  of  guinea-pigs,  causing  death  with  various 
symptoms   only   explainable    on   the   assumption   of   nervous   lesions. 

24  See  Kajjscnbci-'r.  Zcit.  Tiiiimiiiitiit..  1012   (12).  477. 
23  Univ.  of  Ponn.  Mod.  V.uW..   inn.3    (16).  217. 
2«  Trans.  Chicago  Patli.  Soc.   100.3    (5),  207. 


CYToLVs/s  y.v  <;i:m:i{m.   ■  241 

^Microscopically,  the  gaiig'lioii  cells  showed  marked  changes  in  those 
animals  that  survived  the  injection  long  enough.  All  the  results  so 
far  obtained  have  been  with  heterogeneous  serum.-®"*^  Venoms,  par- 
ticularly that  of  cobra,  possess  strong  neurolytic  substances  that  are 
the  chief  toxic  agents  in  most  of  the  venoms  (rattlesnake  venom  ex- 
cepted). 

Thyrolytic  Serum. — There  are  but  few  reports  on  this  sernim,  but 
that  of  Portis  -'  indicates  that  after  removal  of  all  hemolysis  as  a 
factor  there  do  occur  changes,  in  the  nature  of  excessive  absorption 
of  colloid,  and  proliferation  after  the  order  of  that  seen  in  thyroid 
regeneration.  However,  the  clinical  picture  of  thyroidectomy  was 
not  produced  in  any  case,  and  the  anatomic  changes  were  not  great. 
]^y  immunizing  against  nucleoproteins  derived  from  thyroid  tissue, 
Beebe  -"*  has  secured  an  antiserum  to  which  he  ascribes  some  effect 
upon  diseased  thyroids  (exophthalmic  goiter).  MacCallum -"  could 
not  get  a  specific  serum  for  parathyroid  tissue. 

Numerous  reports  may  be  found  indicating  attempts,  with  varying 
success,  to  obtain  serum  toxic  for  other  tissues.  Among  them  may  be 
mentioned  epithcliolysin  -"^  (for  ciliated  epithelium),  spermatotoxin,^'-'^ 
hepatolysin,  cardioUjsin,  splenolysin,  and  syncytiolysm.^°  Special  at- 
tention has  been  given  to  the  production  of  specific  lysins  for  cancer 
cells,  without  definite  success.  (See  Chapter  xvii.)  In  general  it 
can  be  said  that  it  has  }wt  been  found  possible  in  this  way  to  throw 
out  of  function  one  particular  organ,  with  or  without  involvement  of 
other  structures.  The  principles  involved  in  all  these  experiments  are 
the  same,  and  the  results  are  in  no  instance  altogether  satisfactory; 
therefore  no  further  consideration  of  these  special  cytotoxic  serums 
wdll  be  made  here,  the  reader  being  referred  to  other  sources  for  de- 
tails.''^ It  may  be  said,  however,  that  recent  developments  indicate 
that  various  tissues  not  only  contain  proteins  which  exhibit  the  species 
characteristics  of  the  entire  animal,  but  also  other  proteins  or  anti- 
genic radicals  which  are  more  or  less  independent  of  these  and  char- 
acteristic to  a  certain  degree  for  the  tissue  from  which  the  antigen 
was  obtained.     This  being  the  case,  we  cannot  consider  the  problem 

2«a  An  attempt  to  obtain  a  specific  neurotoxin  witli  corpus  striatum  was  un- 
successful.     (Lillian  Moore,  Jour.  Immunol.,  1916    (1),  525.) 

2"  Jour.  Infectious  Diseases,  1904   (1),  127. 

28  Jour.  Amer.  Med.  Assoc,  1906  (46),  484.  Not  corroborated  by  Portis  and 
Bach,  ibid.,  1914   (62),  1884. 

20  Med.  News,  1903   (83),  820. 

2"Ja8ee  Galli-Valerio,  Zcit.  Immunitat.,   1915    (24),  311. 

20b  Taylor  (Jour.  Biol.  Chem.,  1908  (5),  311)  made  the  interestiiif;  oliservation 
that  no  siiermatolytic  serum  could  he  obtained  by  immunizing  with  isolated 
nucleic  acid,  ])r()tamines,  or  ether  extracts  of  sperm,  although  immiuiizing  with 
whole  sperm  jnodiu'cd  active  sera. 

30  Lake,  Jour.  Infect.  Dis.,  1914   (14),  385. 

31  Bioclieniisches  Centralblatt,  1903  (1),  57;5,  rt  sc<j.;  also  see  Sata,  Zicgler's 
Beitr.,  1906    (39),  1;   and  literature  cited  previously. 

16 


242  CHEMlsriiV    OF    the    IMMUMTY    NEACTIOXfi 

of  specific  cytotoxins  a  closed  chapter;  improved  methods  for  sepa- 
rating our  antigens  may  yet  enable  us  to  secure  antibodies  specific  for 
a  single  tissue  or  organ.  (See  Specificity  of  Antigens,  Chap,  vii.) 
However,  by  the  useful  method  of  studying  the  effect  of  cytotoxic 
serum  on  the  growth  of  tissue  cultures  in  vitro,  Lambert  ^-  found  no 
evidence  whatever  of  specificity,  although  there  is  a  non-specific  inhi- 
bition of  growth  by  the  immune  sera. 

32  Jour.  Exp.  Med.,  1914   (19),  277;  also  Walton,  ibid.,  1915   (22),  194. 


CHAPTER    IX 

CHEMICAL  MEANS  OF  DEFENSE  AGAINST  NON- 
ANTIGENIC  POISONS  ' 

Although  the  examples  of  acquired  immunity  against  poisons  of 
known  chemical  composition  are  few  indeed,  nevertheless  the  body 
possesses  means  of  defense  against  many  such  poisons,  which  decrease 
to  greater  or  less  degree  their  harmful  effects.  It  is  to  be  noted,  how- 
ever, that  the  increased  tolerance  to  such  poisons  is  far  less  than  the 
degree  of  tolerance  characteristic  of  immunity  to  true  toxins ;  thus,  in 
arsenic  eaters  the  maximum  observed  tolerance  is  but  three  or  four 
times  the  minimum,  and  less  than  the  certainly  fatal  dose  (Haus- 
mann)  ;  dogs  can  be  made  tolerant  to  only  about  three  times  the  fatal 
dose  of  morphine  (Faust).  Furthermore,  with  many  poisons  of  this 
class  the  tolerance  is  largely  fictitious,  since  in  spite  of  the  absence 
of  acute  symptoms  chronic  poisoning  is  taking  place;  and,  of  course, 
with  many  poisons  no  distinct  increase  of  tolerance  can  be  produced. 
True  immunity,  associated  with  the  production  of  neutralizing  sub- 
stances in  the  blood,  has  as  yet  been  obtained  only  against  substances 
of  protein  nature  or  substances  very  closely  resembling  the  proteins. 
Ehrlich  -  believed  that  simple  toxic  chemicals  are,  like  toxins,  bound 
to  the  cells  by  special  receptors,  cJienioreceptors,  which,  in  view  of 
their  simpler  function  may  be  assumed  to  be  simpler  than  the  re- 
ceptors for  toxins.  They  seem  to  be  more  firmly  fixed  to  the  cells, 
and  being,  therefore,  less  easily  discharged  than  bacterial  receptors 
no  free  antibodies  are  produced  by  immunization.  To  be  sure,  there 
have  been  observations  interpreted  as  evidence  of  immunity  to  large 
molecular  complexes,  especially  such  as  lipoids  and  glucosides,  but 
as  yet  the  positive  establishment  of  the  formation  of  antibodies  by  re- 
action to  non-protein  antigens  has  not  been  accomplished.  It  must  be 
taken  into  consideration,  however,  that  various  chemical  substances  in- 
troduced into  the  blood  or  tissues  of  an  animal,  may  form  compounds 
with  the  animal's  proteins  which  behave  like  foreign  proteins,  to  which 
the  animal  reacts  by  becoming  hypei*sensitive ;  in  this  way  are  ex- 
plained the  instances  of  idiosyncrasy,  with  reactions  of  anaphylactic 
charapter,  which  are  sometimes  shown  with  iodoform,  antipyrine,  sal- 
varsan,  and  other  substances.     (See  Chapter  vii.) 

Studies  on  bacterial  immunity  and  allied  topics  have  as  yet  shown 

1  Bibliography  by  Haiismann,  Ergebnisse  Physiol.,   1907    (6),  58. 

2  Beitriige  z.  exp.  Path.  u.  Chem.,  Leipzig,  1909,  p.  189. 

243 


244  DEFEX^E-  AOAIX^T    XOXAyTWnXW    POISOXS 

notliiiig  to  explain  the  acquirement  of  tolerance  to  morphine,  alcohol, 
arsenic,  and  other  similar  poisons.  A  few  observers  have  claimed  that 
the  serum  of  animals  innnunized  to  morphine  will  neutralize  to  some 
degree  the  toxic  effects  of  mor])hine,  but  these  results  have  not  been 
generally  substantiated.  Others  have  claimed  that  increased  oxida- 
tive powers  are  developed  under  the  stimulation  of  the  poison  which 
permits  of  its  more  rapid  destruction,  especially  in  the  liver,  but  the 
experimental  support  of  this  hypothesis  is  slight.  Still  another  idea 
is  that,  at  least  in  the  case  of  morphine,  decomposition  products  are 
produced,  and  accumulate  in  the  body,  that  neutralize  physiologically 
to  some  extent  the  morphine  itself;  this  hypothesis  can  scarcely  be 
applied  to  arsenic  and  alcohol  tolerance.-^  It  has  been  found  that 
in  animals  habituated  to  morphine  there  is  an  increased  power  to  de- 
stroy morphine,  but,  nevertheless,  the  blood  of  such  animals  still  con- 
tains quantities  of  morphine  toxic  for  normal  animals,  so  there  must 
be  a  certain  refractoriness  or  cellular  immunity  in  addition  (Riib- 
samen).  Schweisheimer  ^^  has  shown  that  when  chronic  alcoholics 
and  total  abstainers  are  given  equal  quantities  of  alcohol,  the  alcohol 
content  of  the  blood  reaches  a  higher  level,  and  persists  for  a  longer 
time  at  a  high  level,  in  the  abstainers.  Apparently  the  alcohol- 
halntuated  organism  can  destroy  alcohol  more  readily,  presumably 
through  more  rapid  oxidation.^"  However,  other  factors  are  involved 
in  alcohol  tolerance,  for  with  equal  quantities  of  alcohol  in  the  blood 
the  abstainers  show  a  more  marked  intoxication  than  the  habitual 
drinker.  So,  too,  in  morphine  tolerance  any  general  resistance  through 
augmented  oxidation  seems  inadequate  in  view  of  the  specific  increase 
in  the  tolerance  of  the  respirator^'  center  observed  in  this  condition.^*" 
Also  we  find  that  tolerance  to  one  drug  may  be  accompanied  by  toler- 
ance to  other  drugs  exerting  similar  physiological  action. ^"^ 

It  is  possible,  also,  that  the  cell  constituents  with  which  the  poisons 
ordinarily  combine  are  produced  in  increased  amounts  under  the 
stimulus  of  the  poison,  just  as  they  are  in  the  case  of  immunization 
with  toxins,  with  the  difference  that  the  combining  substances  are 
not  thrown  off  into  the  blood.  For  example,  it  has  been  claimed  that 
arsenic  is  ordinarily  combined  and  held  in  the  liver  by  a  nucl(M)])r()- 
tein,  and  the  suggestion  has  been  made  that  in  arsenic  habiturs  this 
nucleoprotein  is  increased  in  amount.  Again,  saponin  seems  to  act 
upon  the  cholesterol  of  the  red  corpuscles,  and  Robert  observed   in- 

3  Concernin}?  immunity  ajiainst  moi-iiliiiio  soo  Faust,  Arcli.  oxp.  Tatli.  u.  I'liarm.. 
1000  (44),  217;  Cloctta,  ihid.,  lOO;^  (r)0).  4;"):^;  Kiilisauu'ii.  ihiiL.  l!tOS  ( .V.l) .  -IIT  ; 
Alliiiiicsc,  Arcli.  ital.  bio].,  1010  (5.3),  430 ;  Lan<j;<'r  ( licroiii  tohMaiUH' i .  IliorluMn. 
Zclt..  1012   (4.5).  221:  Valoiiti.  Arcli.  exp.  Patli.,  1014    (75).  4:!7. 

■■iaDout.  Arch,  kliii.  :\Icil..   101:?    (100).   271 

3b  Soo  also  Villi/,  and  Dictricli,  I'.iocJu'ui.  Zcit..  1015  ((iS),  US.  .1.  Ilirsrh. 
ibid.,  1010   (77),  120. 

3<-Van   Doiifrcn.   Arcli.  {Xos.  Physiol.,   1015    (1()2),  54. 

3d  ScKi  flyers.  Jour.  Pharmacol.,  lOlfi  (8),  417.  TTowpvor.  Bilicrfcld  tiiul^^  mor- 
phine tolerance  to  he  specilic    (  Biochem.  Zoit.,   101(5    (77),  2S;l ) . 


^0X-A.\TI(1E\1C  /'O/.S'O.V.S'  245 

creased  resistance  to  the  action  of  saponin  cxliibited  by  tlie  serum  of 
immunized  animals,  which  he  attributes  to  an  increased  amount  of 
cholesterol,  })erhaps  liberated  by  the  corpuscles  decomposed  by  the 
injected  poison,  or  perhaps  produced  in  excess  by  the  tissues.  Wohlge- 
muth ^  has  also  suggested  that  in  the  ease  of  poisoning  with  large 
amounts  of  substances  which  combine  with  glycuronic  acid  (e.  g., 
lysol),  excessive  quantities  of  this  substance  are  formed  by  the  cells 
and  excreted  into  the  blood,  where  they  neutralize  the  poisons  in  much 
the  same  manner  as  the  antitoxins  neutralize  toxins. 

But  besides  these  scanty  examples  of  tolerance  to  poisons,  the  body 
possesses  a  number  of  methods  for  opposing  many  other  poisons  with 
more  or  less  success;  and,  poisons  invariably  acting  chemically,  the 
defenses  are  in  turn  largely  chemical.  We  have  elsewhere  referred  to 
the  destructive  action  of  the  enzymes  of  the  digestive  tract  upon  bac- 
terial and  similar  poisons;  this  means  of  defense  cannot  apply  to 
non-protein  chemical  substances  except  possibly  glucosides  and  toxic 
lipoids.  But  the  acidity  of  the  gastric  juice,  the  alkalinity  of  the 
bile  and  pancreatic  juice,  and  the  precipitating  effect  of  the  hydrogen 
sulphide  formed  in  intestinal  putrefaction  are  all  factors  that  help 
to  neutralize  or  prevent  the  absorption  of  certain  poisons,  their  total 
efficiency,  however,  being  on  the  whole  very  slight.  After  absorption 
of  a  poison  a  large  series  of  chemical  reactions  and  physiological 
processes  is  brought  into  playj  and  there  are  few  poisons  indeed  whose 
harmful  influence  is  not  more  or  less  decreased  by  these  means. 
Robert "'  classifies  these  protective  processes  as  follows : 

1.  Rapid  elimination,  either  before  absorption  by  means  of  diar- 
rhea and  vomiting,  or  by  the  same  means  after  absorption  in  case 
the  poisons  are  excreted  into  the  digestive  tract  (e.  g.,  morphine, 
venoms,  antimony,  and  many  other  metals).  ^Nlany  poisons  are  very 
rapidly  eliminated  bj^  other  routes  {e.  g.,  anesthetics,  curare),  in  some 
instances  causing  harm,  particularly  to  the  eliminating  organ  {e.  g., 
kidneys  in  phenol  poisoning,  intestines  in  ricin  poisoning).  The 
routes  and  conditions  of  elimination  of  poisons  have  been  fully  dis- 
cussed by  Lewin.*' 

2.  Deposition  and  Fixation  in  Single  Organs  or  Tissues. — In  this 
i-espect  the  liver  is  especially  important,  probably  because  of  its  loca- 
tion and  function  as  a  filter  for  all  the  blood  coming  fresh  from  the 
alimentary  tract.'  The  manner  and  means  by  which  this  fixation  is 
brought  about  are  unknown.  It  is  possible  that  the  power  of  the 
tissues  to  bind  poisons  may  become  increa.sed  by  repeated  doses,  lead- 

4  Biochem.  Zeitsehr.,  1906    (1).  1.34. 

5  "Lelnlnicli  der  Intoxikationon."  Stutt;jart. 

6  Dent.  med.  Woch.,  inoo  (32),  10!):  see  also  Mendol  ct  <il..  Amor.  .lour.  Phvsiol., 
1004    (11).  .5;   1906    (16),  147  and  152. 

"  Concerning  the  detoxicatinsr  function  of  tlie  liver  see  Woronzow.  Dissertation, 
Dorpat.  1910:  Rothberger  and  Winter])erg,  Arcli.  intcrnat.  rimrniacodvn.,  1905 
(15),  339. 


246  DEFENSE    AGAINST    NON-ANTIGEMC    POISONS 

ing  to  "specific  acquired  tolerance."^  According-  to  Slowtzoff  * 
arsenic  is  fixed  by  the  nucleus  in  a  very  firm  combination ;  ^°  mercury 
by  globulins  in  a  less  stable  combination;  copper  by  the  nucleins,  but 
less  firmly  than  the  arsenic.  Other  poisons,  chiefly  alkaloids,  are 
probably  combined  with  bile  acids.  Possibly  some  poisons  combine 
with  glycogen.  These  compounds  are  but  slowly  broken  up,  and  thus 
the  poison  reaches  the  more  susceptible  and  more  important  tissues 
in  a  relatively  diluted  condition.  The  bones  seem  to  hold  in  harmless 
form  poisonous  fluorides,  and  to  less  extent  arsenic,  barium,  and 
tungsten,  which  persist  in  the  bones  for  a  great  length  of  time.  Leu- 
cocytes are  possibly  important  binders  of  poisons,  perhaps  through 
combination  with  their  nucleins,^ ^  but  storage  in  these  labile  cells 
is  necessarily  of  relatively  brief  duration.  ]\Iany  poisons  combine 
with  the  inorganic  constituents  of  the  tissues ;  e.  g.,  barium  and  various 
aromatic  substances  with  SO4 ;  silver  with  CI,  etc. 

3.  Combination  with  substances  formed  or  contained  in  the  tissues ; 
the  resulting  substance  being  less  toxic  than  the  poison  alone.  Under 
this  heading  may  be  included  both  chemical  combination  and  physical 
absorption  or  solution,  such  as  the  deviation  of  the  lipoid-solu^e  nar- 
cotics from  the  central  nervous  system  by  excessive  tissue  fat'^?  or  by 
fats  therapeuticallv  introduced.^-  -^ 

4.  Chemical  alteration,  with  or  without  subsequent  combination  with 
other  substances,  by  such  means  as  oxidation,  reduction,  hydrolysis, 
and  neutralization.  •".  "- 

5.  Impaired  absorption  should  also  be  considered  as  a  m^ans  of  de- 
fense against  poisons.  This  may  depend  upon  the  injury  to  the  gas- 
tro-intestinal  tract  produced  either  by  the  poison  itself  or  by  some 
independent  pathological  condition.  Cloetta  considers  impaired  ab- 
sorption important  in  acquired  immunity  to  arsenic  (see  below)  and 
it  may  also  modify  the  effects  of  other  poisons.^^ 

The  chemical  reactions  employed  in  defense  against  simple  chemical 
j>oisons  have  been  particularly  considered  by  E.  Fromm,^^  whose  out- 
line is  here  partially  followed,  and  to  which  the  reader  is  referred  for 
bibliography. 

INORGANIC  POISONS 

Metallic  poisons,  such  as  lead,  silver,  mercury,  and  arsenic,  are 
made  insoluble,  particularly  by  forming  compounds  with  proteins  in 

sSantesson,  Skand.  Arch.  Physiol..  1011    (25),  2S. 

9  Hofmeister's  Reitr.,  1001    (1),  281;    1002    (2),  307. 

10  Denied  by  ITeffter  (Arch,  inteinat.  de  PharmaPodyn..  1005  (15),  300).  who 
considers  it  more  a  physico-chemical  ])rocess. 

11  Stessano,  Conipt.  kend.  Acad.  Sci.,  1900    (131),  72. 

12  See  Graham,  .Tour.  Inf.  Dis.,  1011    (8),  147. 

13  V.  Lhota,  Arch,  internal.  ])harniacodyn..  1012    (22),  QA . 

14  "Die  chemischen  R<'hiit/niittel  des  TierkJirpers  hei  Vcr<.Mf(iiii<rcn."  Strasshnrji, 
Karl  Triibner,  lOO:?.  See  also  n'-sunie  hv  Ellin;:cr.  De\it.  mod.  Wocli.,  lilOO  (2(i), 
580. 


INOR'JAMC  POISONS  247 

the  aliuR'Htaiy  tract,  intestinal  walls,  blood,  or  internal  organs;  also 
by  forming  sulphides  with  the  HgS  of  the  intestinal  contents.  Accord- 
ing to  Cloetta  ^^  immunization  against  arsenic  depends  entirely  upon 
a  reduction  of  absorption  in  the  intestine,  for  the  longer  arsenic  is 
taken,  the  less  appears  in  the  urine  and  the  more  appears  in  the 
feces.^"  At  the  same  time  the  resistance  to  arsenic  injected  sub- 
cutaneously  is  not  increased  at  all,  and  no  increase  in  resistance  can 
be  obtained  by  repeated  subcutaneous  injections  of  sublethal  doses. 
There  is,  however,  reason  to  question  the  authenticity  of  the  reputed 
tolerance  of  habitues  to  arsenic  (Joachimoglu).^"^  Antimony  does  not 
produce  tolerance  in  experimental  animals  (Cloetta). ^^  The  manner 
in  which  various  inorganic  ions  antagonize  the  physiological  action  of 
one  another  (e.  g.,  sodium  and  potassium,  calcium  and  magnesium)  is 
still  an  important  problem. ^'^ 

Free  acids  and  alkalies  are  partly  neutralized  by  the  alkaline  and 
acid  contents  of  the  gastro-intestinal  tract,  partly  by  forming  com- 
pounds with  the  proteins,  and  partly  by  the  alkalies  and  carbonic  acid 
of  the  blood  stream.  (See  "Acid  Intoxication,"  Chap,  xviii.)  Phos- 
phorus ^*  and  sulphides  are  oxidized  after  absorption  into  phosphoric 
and  sulphuric  acid,  which  are  in  turn  neutralized  by  the  alkalinity 
of  the  blood  and  tissues.  Lillie  ^^  has  called  attention  to  the  close, 
palisade  arrangement  of  the  nuclei  of  th^  epithelium  lining  the  ali- 
mentary tract,  which  makes  it  necessary '^f&r  all  substances  absorbed 
to  pass  through  the  zone  of  their  active  oxidative  influence,  a  fact 
undoubtedly  of  great  importance  in  the  defense  of  the  body. 

Reduction  of  iodic  acid,  chloric  acid,  hypochlorous  acid,  and  their 
salts  occurs  in  the  bodv,  resulting  in  their  conversion  into  the  much 
less  toxic  iodides  and  chlorides.  TeUurhim  compounds  are  also  re- 
duced and  rendered  insoluble.  This  reaction  occurs  to  some  extent 
in  the  intestines ;  how  much  in  other  organs  is  unknown. 

Methylation,  the  addition'' of  CH^  groups,  is  observed  in  poisoning 
by  tellurium,  which  is>  eliminated  in  'the  breath  as  methyl  telluride, 
and  also  in  the  sweat  and  fecesjT^  Selenium,  pyridine,  and  some  other 
substances  also  combine  with  methane.  The  source  of  the  methane  is 
possibly  in  the  xanthine  molecule. 

Summary. — There  are,  therefore,  three  chief  reactions  used  against 

15  Arch.  exp.  Path.  u.  Pharm.,'1906  (54),  196:  Corresponbl.  Schweizer  Aerzte, 
1911   (41),  737. 

16  Not  accepted  by  Hausmann.  Ergebnisse  Phvsiol.,  1907  (6),  .58:  or  Joachi- 
moglu,  Arch.  exp.  Path.,  1916   (79),  419. 

IT  Arch.  exp.  Path.  u.  Pharm.,  1911    (64),  352. 

17a  See  Osterhout,  Proc.  Phil.  See,  1916    (55),  533. 

18  Increased  tolerance  to  phosphorus  may  be  obtained  by  repeated  small  doses, 
but  it  lasts  only  while  the  poison  is  beinor  given  continuously  (Oppol,  Ziegler's 
Beitr.,  1910  (49),  543).  Accompanying  the  tolerance  are  structural  changes  in 
the  liver  cells  to  which  are  ascribed  some  significance  bv  Oppel. 

loAmer.  Jour.  Physiol.,  1902   (7),  412. 

20  See  Mead  and  Gies,  Amer.  Jour.  Physiol.,  1901  (5),  105.  Caffein  niav  be 
demethylated  in  the  liver,  Kotake,  Zeit., 'physiol.  Chem.,   1908    (57),   378. 


248  DfJFEXSE    AGAINST    NON-ANTIOENW    POISONS 

inorganic  i)ois()ns  in  the  body,  oxidation,  reduction,  and  splitting  off 
of  tvater;  neutralization  of  acids  or  alkalies  and  the  formation  of  al- 
buminates and  sulphides  being  included  under  the  last  heading,  since 
in  these  reactions  the  splitting  off  of  water  is  an  essential  step. 

ORGANIC  POISONS 

In  the  case  of  organic  poisons  an  equally  small  number  of  primary 
reactions  is  em])loyed  in  their  detoxication,  but  in  more  complicated 
manners  and  condjinations  corresponding  with  the  complexity  of 
organic  compounds. 

Oxidation,  which  has  already  been  mentioned  as  a  means  of  de- 
struction of  bacterial  toxins,  is  naturally  one  of  the  most  effective 
agents  in  the  destruction  of  simpler  organic  substances,  since  the 
ordinary  decomposition  of  all  organic  food-stuifs  is  through  oxidation. 
There  are  numbers  of  specific  examples  of  the  conversion  of  a  poisonous 
into  a  less  poisonous  or  non-poisonous  substance  by  oxidation.  All 
acids  of  the  fatty  acid  series  are  oxidized  vigorouslj^  in  the  body, 
eventually  into  COg  and  H^O ;  and  occasionally  pathologically  pro- 
duced oxalic,  acetic  and  lactic  acids  are  destroyed  in  this  way.  The 
liver  contains  an  oxidase  destroying  alcohol,  which  is  not  increased  in 
the  livers  of  animals  made  tolerant  to  alcohol  (J.  Hirsch).-^  Uric 
acid  is  oxidized  vigorously  by  many  organs,  as  are  other  members  of 
the  purine  series,  such  as  caffeine  and  theobromine.  Presumably  oxi- 
dation of  organic  poisons  as  well  as  of  food-stuffs  is  brought  about  by 
the  oxidizing  enz,ymes  of  the  cells,  as  shown  by  Ehrlich's  indophenol 
reaction,  which  consists  of  the  oxidation  of  paraphenylene  diamine 
and  a-naphthol,  with  a  resulting  synthesis.  This  reaction  is  said  by 
Lillie  "  to  occur  principally  in  and  about  the  cell  nuclei  or  cell  mem- 
branes. 

Combination,  with  or  without  Preliminary  Oxidation, — Oxida- 
tion is  also  an  essential  preliminar.y  step  to  many  of  the  protecting  com- 
binations, in  which  a  cell  constituent  is  united  to  an  organic  poison. 
The  most  important  of  these  combining  substances  are : 

1.  Sulphuric  Acid. — One  of  the  earliest  and  most  impoi'tant  observa- 
tions on  tlie  jjrotective  action  of  sulphuric  acid  was  made  by  Baumann 
and  Herter,--  who  showed  that  phenol  is  eliminate*!  as  a  potassium 
salt  of  the  sulphuric  acid  derivative,  as  follows: 

('„II,()Tr   +   TTO-S(\t\  =  r„Tl,p-SO,l\    +    Il,(). 

a  reaction  that  has  been  put  to  practical  use  in  treating  phenol  jioison- 
ing.  As  phenol  and  cresols  are  produced  constantly  in  intestinal  de- 
composition, this  reaction  is  uiuloubtedly  of  great  service,  since  the  salt 
foi-med  is  relatively  hariidcss.     Indole  and  skatolc  are  similarly  de- 

21  Bioclicin.  /('it.,   line.    (771.  129. 

22  Zi-it.  i)liysi()l.  Cliciii..  1S77    (1).  247. 


ORGANIC  POISONS  249 

toxicatetl  by  being  converted  into  corresponding  salts,  but  only  after 
a  preliminary  oxidation  into  indoxyl  and  skatoxyl,  according  to  the 
following  reaction: 


CM 

C(on) 

/-^ 

/\. 

Coll,     CII  +  0  = 

C'„H,     CH. 

\  / 

\   / 

NH 

Nil 

(indole) 

(indoxyl) 

C(OH) 

'  C— 0— SO.OK 

/^ 

/\v 

CJI,     (  11   +   HO— SOjOK 

=  CH,     CH  +  H..0. 

\   / 

\   / 

KII 

NH 

(indoxyl) 

(indican) 

A  host  of  other  aromatic  organic  substances  are  similarly  combined 
with  sulphuric  acid,-^  with  or  without  preliminary  oxidation,  includ- 
ing all  substances  resembling  phenol  or  which  through  oxidation  are 
changed  into  phenols,  such  as  cresol,  thymol,  anilin,  naphthalin,  pyro- 
gallol,  and  tannin.  By  this  means  a  poisonous  substance  is  converted 
into  a  relatively  harmless  one,  which  is  readily  soluble  and  rapidlj^ 
eliminated. 

2.  Glycuronic  acid  occupies  the  same  position  as  sulphuric  acid,  com- 
bining particularly  with  naphthol,  thymol,  camphor,  chloral  hydrate, 
and  butyl  chloral.  Sometimes  a  substance  may  appear  in  the  urine 
combined  in  part  with  sulphuric,  in  part  with  glycuronic  acid,  show- 
ing the  similarity  of  their  function.  Apparently  when  there  is  not 
sufficient  sulphuric  acid  in  the  body  to  combine  with  all  the  poison, 
the  excess  unites  with  glycuronic  acid,-*  although  combination  between 
glycuronic  acid  and  the  aromatic  substance  begins  to  occur  before  all 
the  sulphuric  acid  is  exhausted.-''  Glycuronic  acid  represents  merely 
a  first  step  in  the  oxidation  of  glucose,  as  follows : 

OHC-(CHOH),-CH,OH  +  0,  =  OHC- (CHOH),-COOH  +  H.O. 
(glucose)  (glycuronic   acid) 

This  oxidation  occurs  after  the  aldehyde  group  of  the  glucose  has 
been  combined  by  some  other  substance ;  hence  the  aldehyde  group 
escapes  oxidation,  although  ordinarily  more  easily  oxidized  than  the 
alcohol  group. 

Just  as  with  the  addition  of  sulphuric  acid,  oxidation  may  be  a 
preliminary  step  to  the  addition  of  glycuronic  acid :  e.  g.,  naphthalin 
is  oxidized  into  a-naphthol,  before  uniting  to  glycuronic  acid,  as  fol- 
lows : 

23  See  Hammarsten's  Text-book    (fourth  American  ed. ),  p.  542. 

24  See  Austin  and  Barron,  Boston  ]Med.  and  Surg.  Jour.,.  190,5  (152),  260. 
Wohlgemuth  has  observed  a  case  in  which  all  the  sulphuric  acid  of  the  urine  was 
in  organic  combination    (Berl.  klin.  Woch..  lOOG   (43).  508). 

25  See  Salkowski,  Zeit.  physiol.  Chem..  1004   (42),  230. 


250  DEFENSE    AGAINST    NON-ANTIGENIC    POISONS 

U     11 

A'=C\        H  H    H 

HCC             J>C—C^  /C=C\          OH 

V  —  Cf           >CH  +  0  =  HCC           \C  —  Ca 

H        V  =  r^  V  — cf          )CH 

H    H  H         \C=C^ 

H     H 

(iiaphtlialiii)  (a-iiaplitliol) 

The  same  is  the  case  with  mam-  camphors  and  terpenes.  Reduction 
may  be  the  preliminary  step,  as  with  chloral  hydrate,  which  is  first 
reduced  to  trichlor-ethyl-alcohol.  In  still  other  cases  splitting  off  of 
w^ater  is  the  chief  preliminary  step. 

3.  Glycocoll  is  one  of  the  longest  known  combining  substances,  the 
observation  of  the  combination  of  glycocoll  with  benzoic  acid  to  form 
hippuric  acid  being  the  first  proof  of  synthesis  in  the  animal  body  dis- 
covered by  Wohler  (1824).     The  reaction  is  as  follows: 

C„H,,COOH  +  H,N-CHo-COOH  =  aH-,CO  —  HN-CH„-CnOH. 
(benzoic   acid)        (glycocoll)  (hippuric   acid) 

A  special  enzyme  has  been  found  in  kidney  substance  which  can  bring 
about  this  reaction  outside  the  body.  Normally  this  enzyme  occurs 
chiefly  in  the  kidney  but  may  also  occur  in  other  organs.  ]\Iany  other 
aromatic  compounds  also  combine  with  glycocoll  before  elimination, 
e.  g.,  salicylic  acid.  Some  are  first  altered  to  a  suitable  form  by 
oxidation ;  e.  g.,  toluene  is  oxidized  to  benzoic  acid,  xylene  to  toluic 
acid,  nitro-benzaldehyde  to  nitro-benzoic  acid.  INIany  of  the  sub- 
stances that  can  be  made  to  combine  with  glycocoll  in  the  body  are  of 
such  a  foreign  nature  that  they  never  could  need  neutralization  under 
any  other  than  experimental  conditions,  but  here,  as  with  the  sul- 
phuric and  glyeuronic  acid  reactions,  combination  occui*s  whenever  a 
suitable  substance  is  present  in  the  blood,  glycocoll  always  being  abun- 
dant as  a  cleavage  product  of  the  proteins. 

4.  Urea  may  also  be  a  means  of  defense,  forming  salts  with  organic 
acids  which  are  rapidly  eliminated  ;  e.  g.,  amido-benzoic  acid  and  nitro- 
hippuric  acid. 

5.  Methane. — IMothylation,  which  occurs  also  with  tellurium,  is 
observed  on  administration  of  pyridine,  as  shown  by  the  following 
equation : 


IT     H 

H     H 

C— C 

C— 0       CIT, 

//       W 

//    ^  y 

TIC         K  +   CH,  +  0  - 

=  BC         X 

\       / 

\       /  \ 

c=c 

C=C       OH 

H     H 

H   n 

(pyridine) 

This  reaction  is  of  special  importance,  because  many  alkaloids  contain 
a  pyridine  group ;  and  the  resulting  methyl  compound  may  be  less 
toxic  than  the  original  alkaloid — e.  g.,  methyl  mori)hine. 


OROANIC  POISONS  251 

6.  Sulphur  si)lit  off  from  proteins  may  coml)ine  with  CNII  and  CNK. 
oonvertinp:  them  into  the  much  less  toxic  sulphocyanides.-"' 

7.  Bile  Acids. — All  tlie  above-mentioned  reactions  are  protective 
largely  beeaiise  the  substances  formed  are  solultie  and  rapidly  elim- 
inated, as  well  as  being  less  toxic  than  the  original  poison.  Com- 
pounds of  many  poisons  are  formed  with  bile  acids  which  are  insoluble, 
and  therefore  only  slowly  dissolve  or  decompose,  thus  protecting  the 
body  from  overwlielming  doses  of  the  poison.  Sncli  eoiu])ounds  are 
fonned,  not  only  with  inorganic  ])oisons,  but  also  with  alkaloids,  espe- 
cially strychnin,  brucin,  and  (|uiiiin.  They  are  then  deposited  in  the 
liver,  to  be  slowly  dissolved  and  eliminated. 

(Occasionally  acetic  acid  and  cijsteine  have  been  observed  to  act  as 
<'ombining  substances.  Calcium  may  be  considered  a  defensive  agent 
against  certain  poisons  [oxalic  and  citric  acids}  with  which  it  forms 
insoluble  compounds,  although  it  is  probable  that  the  toxicity  of  oxa- 
lates dejiends  largely  upon  their  robbing  the  cells  of  calcium.-') 

Neutralization  of  organic  acids  entering  the  body  or  formed  in 
metabolism  is  accomplished  by  the  sodium  carbonate  of  the  blood 
W'hen  in  small  amounts;  if  excessive  in  quantity  (e.  g.,  diabetic  coma), 
a  portion  is  combined  with  ammonia  and  appears  as  an  ammonium 
salt  in  the  urine.  Magnesium  and  calcium  salts  may  also  help  in  the 
neutralization,  probably  at  the  expense  of  the  bone  tissue.-^  (See 
^'Acid  Intoxication,"  Chap,  xviii.) 

Dehydration,  wdiich  plays  a  prominent  part  in  a  number  of  the 
above-mentioned  syntheses,  is  particularly  important  in  the  change 
of  ammonium  carbonate  into  urea : 

XH,— 0  NH„ 

"^('0  =       ^CO  +  2H2O. 
XH,— 0  XH„ 

As  ammonium  salts  of  all  sorts  are  very  toxic,  especially  hemolytic, 
w^hile  urea  is  not,  this  process  is  probably  one  of  the  most  important 
detoxicating  reactions  of  the  body  because  of  the  great  amount  of 
ammonium  compounds  thaf^s  constantly  being  formed  in  nitrogenous 
metabolism. 

Summary. — As  Fromm  points  out,  the  variety  of  reactions  and  the 
variety  of  defensive  substances  are  both  remarkably  small  in  num- 
ber. The  reactions  are :  oxidation  and  reduction,  hydration  and  de- 
hydration, and  perhaps  simple  addition  (methylation).  The  chief 
known  protective  substances  are  the  alkalies  of  the  blood,  proteins, 
hydrogen  sulphide,  sulphuric  acid,  glycocoll,  urea,  cysteine,  bile  acids, 

2G  See  ;Nreurice,  Arch.  int.  Pliarmaeodyn.,  IflOO   (7),  11. 

27  See  Robertson  and  Burnett,  Jour.  Pharmacol.,  1012   (.3),  6.3.5. 

28  In  this  connection  it  may  l)e  mentionerl  that  the  bactericidal  power  of  the 
blood  is  increased  if  the  blood  is  more  alkaline,  decreased  if  it  is  less  alkaline, 
than  usual. 


252  DEFEXSE    AGAINST    NOX-AyTIGEXIC    POISONS 

glj'curoiiic  acid,  and  acetic  acid.  .1/^  these  substances  are  normally 
present  in  the  body,  and  none  of  them  is  specific  against  any  one 
poison,  but  each  combines  with  several  poisons.  This  last  fact  is 
interesting  in  comparison  with  the  highly  specific  nature  of  the  im- 
mune substances  against  bacteria  and  their  products. 

As  far  as  we  know,  no  particular  increase  in  the  neutralizing  sub- 
stances results  from  the  administration  of  inorganic  or  organic 
poisons.  The  body  does  not  appear  to  produce  any  excessive  amounts 
of  sulphuric  acid  in  carbolic-acid  poisoning,  or  of  glycocoll  when 
benzoic  acid  is  administered.  Both  substances  are  present  in  the 
body  normally,  and  as  much  as  is  available  combines  with  the  poison ; 
if  there  is  not  enough,  the  remaining  poison  combines  with  something 
else,  or  goes  uncombined.  In  other  words,  the  neutralizing  substances 
described  above  do  not  appear  to  be  the  result  of  any  special  adapta- 
tion to  meet  a  pathological  condition.  They  are  present  in  the  body 
as  a  result  of  normal  metabolism ;  they  have  an  affinity  for  various 
chemical  substances,  some  of  w^hicli  happen  to  be  poisons ;  if  these 
poisons  happen  to  enter  the  body,  they  may  be  combined  and  neutral- 
ized to  some  extent,  but,  as  a  rule,  very  incompletely.  There  appears 
to  be  no  elaborate  process  of  defense  against  the  chemically  simple 
poisons,  such  as  seems  to  be  called  into  action  by  bacterial  infection, 
and  hence  a  degree  of  resistance  or  immunity  similar  to  that  present 
after  an  attack  of  scarlet  fever  or  smallpox  does  not  exist  for  strychnin 
or  phosphorus. 

It  is  also  of  interest  to  consider  that  unicellular  organisms  may 
show  a  marked  capacity  to  increase  their  resistance  to  poisons,  as 
shown  especially  by  Ehrlich's  studies  on  trypanosomes,  which  readily 
become  immune  to  various  trypanocidal  drugs,  including  arsenic 
compounds,  and  which  transmit  this  acquired  immunity  through  suc- 
ceeding generations.  Yeasts  and  bacteria  can  also  exhibit  increased 
tolerance  to  antiseptics,  and  Effront  found  that  yeasts  owe  their  aug- 
mented tolerance  to  fluorides  to  an  increased  content  of  calcium, 
which  precipitates  the  fluoride  which  enters  the  cells ;  this  tolerance  is 
also  transmitted  to  new  generations  of  yeasts.  The  acquired  tolerance 
is  specific  in  all  these  cases,  and  may,  indeed,  be  accompanied  by 
a  decreased  resistance  to  other  poisons ;  thus,  protozoa  acclimated  to 
alcohol  may  be  more  susceptible  to  other  chemicals.-"  Paramecia 
made  immune  to  antimony  are  not  immune  to  arsenic,  and  this  specific 
immunity  is  transmitted  to  succeeding  generations   (Neuhaus).^" 

20  Daniel,  .Jour.  Exper.  Zool.,  1000    (fi),  .571. 

30  Arch.  Internat.  Pliarmacoydn.,  1910   (20),  .303. 


CHAPTER    X 
INFLAMMATION,'   REGENERATION,  GROWTH 

Although  morphological  alterations  are  prominent  features  of  the 
reaction  of  the  tissues  to  local  injury  and  infection,  yet  at  the  bottom 
the  processes  of  intlamniation  are  brought  about  by  and  result  in 
chemical  alterations.  The  causes  of  inflammation  are  in  nearly  all 
cases  chemically  active  substances,  but  for  the  most  part  their  nature 
is  too  little  known  to  permit  of  speculation  as  to  what  chemical  char- 
acteristic or  characteristics  a  substance  must  possess  to  exhibit  the 
power  of  causing-  an  inflammatory  reaction.  Even  in  the  case  of  in- 
flammation due  to  mechanical,  thermal,  and  electrical  injuries,  it 
seems  probable  that  most  of  the  features  of  the  inflammatory  reaction 
are  brought  about  by  the  action  of  chemical  substances  produced  by 
alterations  in  the  tissue  constituents  at  the  point  of  injury,-  for  tissue 
proteins  that  have  been  altered  in  necrosis  are  chemotactic,-'^  as  also 
are  extracts  of  tissues. 

The  essential  features  of  inflammation,  namely,  local  hyperemia 
and  related  vascular  disturbances,  exudation  of  plasma,  migration  of 
leucocytes  and  their  phagocytic  action,  all  may  be  caused  by  the  action 
of  chemical  substances  upon  the  vessels  and  leucocytes.  Active  hy- 
peremia in  the  case  of  inflammation  is  due  to  stimulation  of  the 
vasodilator  nerves  or  paralysis  of  the  vaso-eonstrictors,  or  direct  par- 
alysis of  the  muscular  fibers  of  the  arterioles ;  these  may  result  from 
mechanical,  thermal,  or  electrical  stimuli,  but  in  local  infection  the 
cause  is  usually  chemical  products  of  bacterial  growth  or  of  tissue 
disintegration.  The  escape  of  hlood  plasma  (inflammatory  edema) 
appears  to  depend  upon  a  number  of  factors   (discussed  more  fully 

1  For  extensive  reviews  and  bibliography  see  Adanii,  in  Allbutt's  Rvstem  of 
Medicine;  reprinted  also  as  a  monograph.  "TnflamTnation,"  1000;  also  Opie.  Arch. 
Int.  ^ted..  1910  (.5),  541.  Some  interesting  ideas  are  advanced  by  Klemensiewicz, 
"Die  Entziindung,"  G.  Fischer,  .Tena.  li'OS. 

2Schlaepfer  (Zeit.  exp.  Path..  1010  fS).  ISl)  finds  tliat  tlic  reduction  of 
methylene  blue  is  decreased  in  inflammatory  areas,  and  advances  the  hypothesis 
that  inflammatory  stimulants  are  oxidation  stimulants,  inflammation  occurring 
only  when  the  amount  of  oxidation  aroused  l)y  the  stimulant  is  insufficient.  In 
accord  with  this  is  the  observation  of  Amberg  (Zeit.  exp.  ^Ted.,  101."}  (2).  10) 
that  substances  facilitating  oxidation  reduce  inflammatory  reactions.  (See  also 
Woolley,  Jour.  Amer.  ^Med.,  Assoc,  1014  (6.3),  2270.)  Another  observation  of 
similar  significance  is  that  phagocytosis  is  stimulated  by  H2O2,  and  that  phago- 
cytes react  to  HXC  in  the  same  wav  as  the  respiratorv  center  (Hamburger;  In- 
ternat.  Zeit.  plivs.-chem.  Biol..  101.3  "(2),  24.5-2(54). 

2a  Burger  and  Dold,  Zeit.  Immunitiit..  1014    (21),  .378. 

2,-S.3 


254  INFLAMMATION,  REGENERATION,  GROWTH 

under  "Edema,"  Chap,  xii)  of  which  the  most  important  seem  to  be: 
(1)  injury  to  the  capiHary  M'alls,  produced  largely  by  the  chemical 
causes  or  products  of  the  inflammation-.  (2)  increased  osmotic  pres- 
sure in  the  tissues,  due  to  increased  or  abnormal  formation  of  crystal- 
ioidal  substances  with  high  osmotic  pressure  from  large  molecular 
compounds,  many  of  which  are  colloids  (proteins)  without  apprecia- 
ble osmotic  pressure;  (3)  alterations  in  the  hydration  capacity  of  the 
colloids,  whereby,  tlirough  decrease  in  salts  or  increase  in  acidity, 
thej^  come  to  possess  a  greater  affinity  for  w^ater  (^I.  H.  Fischer). 
By  far  the  most  characteristic  feature  of  inflammation,  however,  is 
the  hehavior  of  the  leucocytes — their  increase  in  number  in  the  blood, 
their  migration  from  the  vessels  and  accumulation  about  the  point 
of  injury,  and  their  engulfing  and  destroying  various  solid  particles, 
such  as  bacteria  and  degenerating  tissue  elements.  These  processes, 
which  seem  to  indicate  something  approaching  independent  volition 
on  the  part  of  the  leucocytes,  may,  however,  be  well  explained  by  ap- 
plication of  known  laws  of  chemistrv'  and  physics,  without  passing 
into  the  realms  of  the  metaphysical.  This  will  be  attempted  under 
the  heading  of : 

AMEBOID  MOTION  AND  PHAGOCYTOSIS 

The  accumulation  of  leucocytes  at  a  given  point  in  the  body  indi- 
cates that  some  means  of  communication  must  exist  between  this 
point  and  the  leucocytes  in  the  circulating  blood.  No  direct  com- 
munication by  the  nervous  system  or  other  stnictural  method  existing, 
the  only  possible  explanation  is  that  the  connnunication  is  through 
the  fluids  of  the  body,  and  depends  upon  changes  in  their  chemical 
composition  or  physical  condition.  As  the  latter  generally  depends 
upon  the  former,  the  communication  is  considered  to  be  accomplished 
by  chemical  agencies,  and  called  chemotaxis. 

CHEMOTAXIS 

Changes  in  the  chemical  composition  of  a  fluid  have  been  shown 
frequently  to  aflPect  the  motion  of  living  organisms  suspended  in  it. 
One  of  the  earliest  observations  was  that  of  Engelmann,^  who  no- 
ticed that  Bacterium  termo  suspended  in  water  tended  to  accumulate 
about  a  bubble  of  oxj^gen  in  the  water.  Pfeffer  *  discovered  that  the 
spermatozoids  of  certain  ferns  were  attracted  powerfully  by  veiy  dilute 
solutions  of  malic  acid,  which  is  contained  in  the  female  sperm  cell, 
indicating  that  the  migration  of  the  sperm  cells  in  the  proper  direc- 
tion depends  on  a  chemical  communication,  and  he  proposed  the  term 
chemotaxis  for  this  phenomenon.  Strong  solutions  of  malic  acid,  on 
the  other  hand,  repelled  spermatozoids.     Cane-sugar  was  found  to  at- 

3  Botanisoho  Zoitiinp,  ISSl    (.30).  441. 

♦Untersuoh.  aiis  dem  Bot.  Institut  in  Tiiliiii;:(>n,  1SS1-1SSS.  Bd.   1   und  2. 


CHEMOTAXIS  255 

tract  tlic  spermato/oids  of  a  certain  I'oliacoous  moss.  In  the  case  of 
the  malic  acid,  it  seems  to  be  the  anion  that  produces  tlie  effect,  since 
salts  of  malic  acid  have  exactly  the  same  property. 

Stahl's  •"'  ex])eriment  with  a  large  jelly-like  Plasmodium  {Aethal- 
twn  scpticu))))  p'rowing  on  hark  in  wet  places,  has  become  classical. 
He  found  that  if  the  plasmodium  was  placed  on  a  moist  surface,  and 
nearby  was  placed  a  drop  of  an  infusion  of  oak  bark,  the  organism 
moved  by  the  process  of  ])rotoplasmic  streaming  toward  and  into  the 
infusion.  If  a  piece  of  oak  bark  was  placed  in  the  water,  plasmodial 
arms  were  sti'ctched  out  to  it  and  the  piece  of  bark  was  soon  com- 
pletely surrounded  by  the  organism.  These  movements  were  found 
to  occur  in  any  direction,  even  exactly  against  the  force  of  gravity. 
Other  substances,  as  acids  or  strong  solutions  of  salt  or  sugar,  were 
found  to  cause  the  phismodium  to  move  away  from  them,  although 
when  sufficiently  dilute  they  exerted  an  attraction.  A  plasmodium 
might,  however,  move  into  a  strong  sugar  solution  if  kept  with  a 
scanty  supply  of  moisture  for  some  time,  and  after  it  had  lived  in 
such  a  strong  solution  (2  per  cent.)  for  some  time,  a  weaker  solution 
or  pure  water  was  as  injurious  as  the  concentrated  sugar  solution 
previously  had  been. 

Temperature  was  also  found  to  exert  a  marked  thermotactic  effect. 
If  a  Plasmodium  was  placed  on  a  filter-paper,  on^  end  of  which  was 
in  water  at  7°,  and  the  other  in  water  at  30°,  it  would  move  toward 
the  warmer  end. 

The  Theory  of  Tropisms. — Ciliated  protozoa,  which  can  move 
about  freely  in  water,  show  very  distinct  reactions  to  stimuli  of  all 
sorts.  The  first  step  in  their  change  of  direction  of  movement  is 
considered  by  many  observers  to  be  an  orientation  of  the  organism 
until  it  is  headed  in  the  axis  along  which  it  is  to  move.  This  is 
ascribed  by  J.  Loeb  **  to  the  existence  of  a  certain  degree  of  equality 
of  irritability  of  sjrmmetrical  parts  of  the  body.  The  stimulant, 
Avhether  it  be  rays  of  light,  or  diffusing  chemicals,  or  heat-waves, 
moves  along  definite  lines,  and  the  organism  receives  at  first  unequal 
stimuli  on  symmetrical  parts  of  the  body,  unless  the  axis  of  the  organ- 
ism is  parallel  to  the  lines  of  motion  of  the  stimulant.  As  long  as  the 
stimulant  acts  on  symmetrical  parts  of  the  body  unequally,  these  parts 
will  react  unequally  until  at  length  the  body  is  swung  into  a  position 
where  the  stimulation  is  equal,  when  it  will  stay  in  this  position  and 
move  along  a  line  parallel  to  the  line  taken  by  the  stimulant.  Not 
only  protozoa,  but  much  higher  forms,  including  some  vertebrates 
are  believed  to  react  in  this  way  to  stimuli — e.  g.,  the  maintenance  bj 
fish  of  a  position  heading  up  stream.  The  above  constitutes  the  so- 
called  'theory  of  tropism,"  and  we  have  such  reactions  to  stimuli  of 
all  sorts,  not  only  chemotropism  and  thermotropism,  but  also  helio- 

sBotanisehe  Zeitunp,  1884   (42),  145  and  161. 

6  Comparative  Physiology  of  the  Brain,  New  York,  1900,  p.  7. 


256  IXFLAMMATION,  NEGENEKATIOX,  GROWTH 

tropism  (reaction  to  light)  ;  geotropism   (to  gravity),  electropism  (to 
electricity),  thigmotropism  (reaction  to  contact),  etc. 

The  work  done  upon  tropisms  applies  particularly  to  ciliated, 
freely  motile  organisms,  and  interests  us  less  in  connection  with 
leucocytes  than  do  the  observations  on  such  forms  as  AmcehaJ  In 
passing  maj'  be  mentioned  the  fhiynwtaxis  or  thigmotropism  (reac- 
tion to  mechanical  stimuli)  shown  by  spermatozoa,  which  explains 
their  apparently  difficult  feat  of  advancing  in  opposition  to  the  cilia 
of  the  epithelial  lining  of  the  female  generative  tract.  It  may  also  be 
noted  that  the  nature  of  reactions  of  organisms  to  various  stimuli  is 
not  constant  for  even  the  same  organism.  Copepods  (minute  crus- 
taceae)  may  be  negatively  heliotropic  in  the  day  and  go  away  from 
the  bright  surface  of  the  water,  whereas  at  night  the  same  animals 
are  positively  heliotropic  and  swarm  to  the  surface  illuminated 
brightly  by  a  lantern.  Variations  in  heliotropism  may,  in  some  cases, 
be  explained  as  due  to  chemical  changes  that  occur  in  the  organism, 
which  explanation  is  made  more  probable  by  J.  Loeb's  experiments, 
Avhich  show  that  change  in  composition  in  the  fluid  in  which  animals 
are  suspended  may  cause  a  complete  reversal  in  their  reaction  to  a 
constant  stimulus.  IMotile  bacteria  seem  to  behave  much  like  ciliated 
protozoa  in  their  reaction  to  stimuli. 

CHEMOTAXIS  OF  LEUCOCYTES  « 

That  leucocytes  come  to  the  site  of  an  infection  because  of  chemical 
substances  produced  by  bacteria  at  this  point,  that  is  to  say,  through 
chemotaxis,  was  first  clearly  pointed  out  by  Leber  ^  in  1879,  who 
likened  the  attraction  of  such  substances  for  leucocytes  to  the  effect 
of  malic  acid  upon  spermatozoids  as  shown  by  Pfeffer.  He  found 
that  in  keratitis,  leucocytes  invaded  the  avascular  cornea  from  the  dis- 
tant vessels,  not  in  an  irregular  manner,  but  all  moved  directly  toward 
the  point  of  infection,  where  they  collected.  As  dead  cultures  of 
staphylococci  produced  a  similar,  although  less  marked,  accumlilation 
of  leucocytes,  he  sought  the  chemotactic  substance  in  their  bodies,  and 
isolated  a  crystalline,  heat-resisting  substance,  phlogosin,  which  at- 
tracted leucocytes  in  animal  tissues.  He  also  observed  that  capillary 
tubes  filled  with  phlogosin  or  with  staphylococci  were  soon  invaded  by 
masses  of  leucocytes. 

Since  Leber's  experiments,  many  other  investigations  have  been 
made  showing  that  chemical  substances  of  many  different  origins  other 
than  Ijacterial  exert  a  chemotactic  influence  on  leucocytes.  Some  sub- 
stances are  indifferent  in  effect,  most  are  positive,  while  some  are  be- 
lieved to  repel  leucocytes;  /.  e.,  are  negatively  chemotactic. 

7  For  full  details  see  .Toiiniii'xs  (Publication  Xo.  10.  Caniotjie  tustituto.  Wash- 
ington, l!tn4:  also  J.  Loch,  "Studies  in  (Icncral   Pliysiolojiv." 

8  Keview  of  literature  on  leucocytes  bv  TTellv,  Krjieb.  allix.  Patliol.,  I!tl4  (17,,,),  1. 
0  Fortschritte  der  :\Ied.,   1888    (6),  4Gf>. 


CHEMOTAXIS  OF  LEUCOCYTES  257 

Negative  Chemotaxis. — Probably  the  substances  that  repel  leuco- 
cytes are  iV-w  in  number;  Kantliack,  indeed,  doubted  the  existence  of 
really  negative  cliemotactic  action  upon  leucocytes.  Verigo  ^°  also 
considoi's  that  as  yet  no  actual  negative  chemotaxis  has  been  satisfac- 
torily demonstrated;  but,  by  analogy  with  the  effects  of  chemicals  on 
amebae,  ciliata,  and  plasmodial  forms,  which  all  siiow  a  decided  nega- 
tive chemotaxis  under  certain  influences,  it  would  seem  most  probable 
that  leucocytes  also  should  be  repelled  as  well  as  attracted  by  chem- 
icals.^^ 

Non=bacterial  Chemotactic  Substances. — One  of  the  earliest  sig- 
nificant studies  of  the  effects  of  non-bacterial  substances  upon  chem- 
otaxis was  made  by  ]\Iassart  and  Bordet,^-  who  showed  that  products 
of  the  disintegration  of  leucocytes  and  other  cells  had  a  strong  posi- 
tive chemotactic  influence.  They  also  corroborated  the  statement  of 
Yaillard  and  Vincent  that  lactic  acid  is  an  actively-  repellant  sub- 
stance, for  they  found  that  tubes  containing  a  pyocyaneus  culture, 
which  ordinarily  became  filled  with  leucocytes  rapidly,  did  not  become 
invaded  at  all  if  lactic  acid  was  also  added  in  a  strength  of  1  :  500, 
although  leucocytes  did  enter  when  the  dilution  was  1  :  1000. 

Gabritchevsky  ^^  studied  the  chemical  influence  of  a  large  number 
of  substances  on  leucocytes  and  divided  them  into  three  groups :  I. 
Substances  exerting  "negative  chemotaxis,"  including  those  that  at- 
tracted only  a  few  leucocytes.^*  II.  Substances  with  "indifferent 
chemotaxis,"  which  attracted  moderate  numbers  of  leucocytes.  III. 
Substances  with  positive  chemotaxis.  If  we  correct  the  groupings 
made  by  Gabritchevsky  we  have  the  following  classification: 

I.  Substances  negatively  chemotactic  or  indifferent : 

(a)  Concentrated  solutions  of  sodium  and  potassium  salts; 
(&)  Lactic  acid  in  all  concentrations;  (c)  quinine  (0.5  per 
cent.)  ;  (d)  alcohol  (10  per  cent.)  ;  (e)  chloroform  in  wa- 
tery solution;  (/)  jequirity  (2  per  cent.,  passed  through 
Chamberland  filter)  ;  (g)  glycerol  (10  per  cent,  to  1  per 
cent.);  (h)  bile;  (i)  B.  cliolerae  gaUinarium. 
II.  Substances  with  feeble  chemotaxis : 

(a)    Distilled   water;    (&)    dilute   solutions   of   sodium   and 

10  Arch.  d.  MM.  exper.,  1001    (13),  .585. 

11  Salomonsen's  observation  (Festskrift  ved  indvielsen  af  Statens  Sorum  In- 
stitut,  Kopenhagen.  1902,  Art.  XII),  that  ciliated  infusoria  when  killed  show 
a  strong  negative  effect  on  other  ciliates,  is  of  much  interest,  particularly  as  it 
seems  to  be  the  opposite  of  tlie  positively  chemotactic  effect  of  dead  upon  living 
leucocytes.  The  negative  reaction  of  different  ciliata  was  specific  for  their  own 
kind  rjuantitatively.  but  not  qualitativelv. 

12  Ann.  d.  I'Inst!  Pasteur,  1891    (5),  417. 

13  Ann.  d.  Tlnst.  Pasteur,  1890   (4),  346. 

1*  Evidently   these   substances   were    not    all    negatively    chemotactic,    but    were 
relatively  sliQ;htly  chemotactic  or  indifferent:  yet  in  the  literature  generally  these 
experiments  Jiave  been  cited  as  indicating  a  negative  chemotactic  influence  of  the 
substances  .studied. 
17 


258  IXFLAMMATIOX,  REGENERATION,  GROWTH 

potassium  salts   (1-0.1  per  cent.);   (c)   phenol;   (d)   anti- 
pyrin;  (e)  phloridzin ;  (/)  papayotin  (in  frog)  ;  (  g)  glyco- 
gen;  (h)   peptone;    (i)    bouillon;    (j)    blood  and  aqueous 
humor;  (k)  carmine. 
III.  Substances  with  strong  positive  chemotaxis : 

(a)    Papayotin    (in  rabbits)  ;    (b)   sterilized  living  cultures 
of  bacteria,  whether  pathogenic  or  non-pathogenic. 

These  results  can  only  be  considered  as  suggestive  and  not  as  accu- 
rate findings,  in  view  of  other  contradictory-  results.  Buchner  ^^ 
obtained  from  the  piwumohacillus  of  Friedlander,  a  protein  which  ex- 
erted a  strong  chemotactic  influence,  thus  showing  the  chemical  na- 
ture of  the  attraction  of  leucocytes  by  bacteria,  and  he  isolated  other 
similar  proteins  from  other  bacteria.  He  also  obtained  a  "glutin- 
casein"  from  grain  which  was  related  chemically  to  the  bacterial  pro- 
teins, and  which  was  equally  chemotactic.  The  metabolic  products 
of  bacteria,  however,  he -found  to  be  negatively  chemotactic.  Alkali 
albuminate  and  hemi-albumose  were  strongly  positive,  but  peptone 
was  not.  GlycocoU  and  leucine  were  found  to  be  chemotactic,  but 
urea,  ammonium  urate,  skatole,  tyrosine,  and  trimethylamine  were 
not.  It  was  also  observed  that  if  the  positively  chemotactic  sub- 
stances were  injected  subcutaneously,  they  produced  general  as  well 
as  local  leucocytosis.  The  products  of  the  action  of  serum  on  bacteria, 
"  anaphylatoxin, "  produce  inflammatory  reactions,  and  probably  are 
important  factors  in  pathology ;  the  products  of  tissue  disintegration 
have  similar  effects.^^'^  Certain  drugs  (notably  quinine,  morphine 
and  chloral)  when  injected  subcutaneously  seem  to  reduce  the  amount 
of  leucocytic  emigration  at  a  point  of  local  injury  (Ikeda).^^'' 

V.  Sicherer  ^®  found  that  chemotaxis  of  leucocytes  may  be  observed 
outside  the  body.  If  a  tube  containing  positively  chemotactic  sub- 
stances (dead  beer-yeast  cells  and  dead  staphylococci  were  the  strong- 
est) is  placed  with  one  end  in  a  leucocyte-containing  exudate,  the  leu- 
cocytes pass  up  into  it  against  gravity. 

Bloch  ^■^  demonstrated  that  carbol-glycerol  extracts  made  from  each 
of  the  different  viscera  and  tissues  exerted  a  positive  chemotaxis,  dis- 
crediting the  statements  of  Goldscheider  and  Jacob  that  onlj-  extracts 
of  hematogenetic  tissues  showed  positive  chemotaxis.  Egg-albumen, 
gelatine,  albumen-peptone,  and  alkali  albuminate  were  also  positive, 
carboliydrates  feebly  so,  and  fat  not  at  all.  ^Metallic  copper,  iron, 
mercury,  and  their  salts  have  also  been  found  to  be  chemotactic  sub- 
stances, but  it  is  very  probable  that  they  act  in  part  through  destroy- 
ing the  tissues  in  their  vicinity,  which  give  rise  to  decomposition-prod- 

15  Berl.  klin.  Wochensehr.,  1890   (27),  1084. 

15a  See  Bold,  Arh.  Patli.  Inst.  Tiibingen.  1J)14    (i1).  :J(). 

15b  .Jour.  Pharniacol.,  litHi   (8),  137. 

10  Cent.  f.  Bakt.,  lS!)i)    (2(i),  300. 

17  Cent.  f.  allf,'.  I'atli.,  IBJIO    (7),  785. 


yON-BACTERIAL  CHEMOTACTIC  SUBSTANCES  259 

nets  having  a  positive  effect.  Adler,^*  however,  found  that  bichloride 
of  mercury  as  dilute  as  1  :  3000  caused  more  leucocytic  invasion  of  a 
piece  of  saturated  older  pith  than  did  even  cultures  of  pyogenic  bac- 
teria.^" 

^Tetehnikoft'  obsemcd  tliat  leucocytes  might,  after  a  time,  be  at- 
tracted toward  substances  that  at  first  seemed  to  repel  them.  If  the 
blood  is  full  of  toxins,  the  subcutaneous  introduction  of  bacteria  does 
not  lead  to  a  local  accumulation  of  leucocytes,  presumably  because  the 
difference  in  chemotaxis  between  the  blood  and  the  tissue  fluids  is  not 
great  enough  to  cause  emigration ;  in  this  connection  should  be  men- 
tioned Pfeffer's  observation  that  B.  termo  in  a  peptone  solution  will 
not  migrate  toward  another  stronger  peptone  solution,  unless  the  lat- 
ter is  at  least  five  times  as  strong  as  the  former.  Leucocytes  will  mi- 
grate freely  toward  substances  that  kill  them;  of  the  bacterial  prod- 
ucts the  toxins  of  pyocyaneus  and  diphtheria  bacilli  being  especially 
destructive  and  causing  typical  karyorrhexjs.-°  Substances  soluble 
in  lipoids  are  said  by  Hamburger  ^^  to  increase  phagocytic  activity 
when  in  extreme  dilutions,  although  stronger  concentrations  are  highly 
toxic  for  leucocytes.  If  an  electric  current  is  passed  through  two  fin- 
gers there  will  be  found  more  leucocytes  in  the  tissues  of  the  cathode' 
finger  than  in  the  anode  finger,  presumably  because  the  OH-ions  in- 
crease ameboid  movement. "^^ 

]\rany  substances  have  been  used  to  increase  the  number  of  leuco- 
cytes in  the  circulating  blood  in  the  hope  of  increasing  resistance  to 
infections,  a  result  that  does  not  seem  to  follow  artificial  leucocytosis 
with  any  recognizable  uniformity.  A  compilation  of  the  literature 
on  this  subject  by  Gehrig  -'^'^  shows  such  contradictor}'  findings  as  to 
indicate  that  most  of  the  recorded  M'ork  is  of  little  value.  He  was 
unable  to  corroborate  the  current  statement  that  antipyretic  drugs 
increase  the  number  of  leucocytes  in  the  blood.  Nucleinic  acid  and 
tissue  extracts  seem  to  increase  circulating  leucocytes  with  considerable 
regularity,  while  witli  thorium-A'  and  benzol  they  can  be  reduced  to 
almost  complete  extinction.  The  behavior  of  inflammatory  processes 
in  animals  thus  deprived  of  available  leucocytes  has  considerable  ex- 
perimental interest.-^'^  If  less  than  1000  leucocytes  per  cubic  mm. 
are  present  in  the  blood,  no  leucocytic  exudate  can  be  produced, ^^"* 
although  the  other  features  of  inflammation  occur  as  usual. 

Relation  of  Cell  Types  to  Migration. — Of  the  leucocytes,  the 
most  actively  affected  by  chemotaxis  is  the  polymorphonuclear  vari- 

is  Festschr.  for  A.  Jacobi,   1000,  Xew  York. 

10  Concernincr  th^  effects  of  iodin  and  iodides  Tij>on  tlio  leueocvtes,  see  Heinz, 
Virohow's  Arch.,  1809    (1.55).  44. 

20  Schiirmaiin,  Cent.  f.  Pathol..  1010    (21).  .3.37. 

21  Arch.  N^erland..   1912    (III,  B),   1.34:   Brit.  Med.  .Jour.,   1016    (1),  37. 
2iaSchwvzer,  Bio<'heni.  Zeit..   1914    (60),  454. 

2ibZeit.  exp.  Path..  1015    (17),  161. 

2ieSee  G.  Rosenow,  Zeit.  exp.  Med..  1914    (3).  42. 

2i<i  Camp  and  Baumgartner,  Jour.  Exp.  Med.,  1915   (22),  174. 


260  IXFLAMMATIOX,  REGENERATION,  GROWTH 

ety,  but  not  all  substances  affect  each  variety  of  leucocyte  in  the  same 
M'ay ;  for  example,  infections  with  most  animal  parasites  result  in  both 
local  and  general  increase  in  the  eosinophilous  forms,  and  similar  ef- 
fects have  been  obtained  by  the  injection  of  extracts  of  animal  para- 
sites. Lymphocytes  are  much  less  active,  presumably  because  they 
contain  less  of  the  mobile  cytoplasm  and  consist  chiefly  of  the  struc- 
turally fixed  nuclear  substance.  Undoubtedly  many  of  the  cells  in  so- 
called  lymphocytic  accumulations  seen  in  certain  conditions,  such  as 
tuberculosis,  are  not  lymphocytes  from  the  blood,  but  are  newly  di- 
vided cells  of  the  tissue.--  The  experimental  evidence  concerning 
lymphocytic  emigration  is  very  contradictory.  Fauconnet  -^  has  found 
that  tuberculin  injections  cause  in  man  general  increase  in  leucocytes, 
but  only  of  the  polymorphonuclear  form.  Long-continued  intoxica- 
tion of  animals,  however,  may  result  in  lymphocytic  increase,  but 
local  introduction  of  the  toxin  leads  to  accumulation  of  polymorpho- 
nuclear cells  and  not  lymphocytes. 

Particularly  significant  is  the  experiment  of  Reckzeh,-*  who  found 
that  in  Ij-mphatic  leukemia,  with  the  lymphocytes  greatly  exceeding 
the  polymorphonuclear  forms  in  the  blood,  the  pus  from  an  acne  pus- 
tule or  from  cantharides  blisters  contains  practically  no  lymphocytes, 
but  is  composed  of  the  usual  polynuclear  forms.  WolfiC  -^  however, 
claims  that  tetanus  and  diphtheria  toxins  produce  lymphocytosis  in 
experimental  animals.  Wlassow  and  Sepp  -'^  state  that  lymphocytes 
are  not  capable  of  ameboid  movement  or  phagocytosis  in  the  body, 
although  after  heating  to  44°  they  may  become  motile  for  a  short 
time. 

Experiments  on  the  nature  of  the  leucocytes  attracted  by  different 
ehemotactic  agents  have  been  made  by  Borissow  -^  and  Adler.-^  Both 
agree  in  stating  that  none  of  the  substances  tested  shows  any  special 
affinity  for  any  single  type  of  leucocytes.  Zieler  -^  seems  to  have  set- 
tled this  matter  positively  by  his  observation  that  in  the  skin  of  rab- 
bits exposed  to  the  Fiiisen  light,  active  migration  of  lymphocytes  takes 
a  prominent  part  in  the  reaction.  General  lymphocytosis  may  be 
produced  by  certain  substances  (pilocarpine,  muscarine,  BaCL)  which 
cause  contraction  of  the  smooth  muscles  and  force  these  cells  out  of 
the  spleen  ( Harvey ),^°  but  such  a  process  has  no  relation  to  chemo- 
taxis.     It  is  notorious  that  infections  with  animal  parasites  cause  both 

22  See  r^siim<5  bv  Papponlioini.  Folia  Ilematol.,  lOO.l   (2).  Sla:   1006   (3),  120. 

23Deut.  Arch.  klin.  Mod..  1004   (82),  167. 

24Zeit.  f.  klin.  Med.,  1003    (50),  51. 

25TVrl.  klin.  Wocli.,  1004    (41),  1273. 

2«  Vircliow's  Arch.,  1004    (176),  185. 

27Ziep;ler's  Beitriio;e,   1804    (16),   432. 

28  Festschrift  f.   A.  .Tacobi,  New  York,   1000. 

29  Cent.  f.   Pathol.,   1007    (18),  280. 

•iojour.  of  Phvsiol.,  1906  (35),  115;  see  also  Pvoiis,  Jour.  Fxper.  Med.,  1008 
(10),  238. 


TEERMorAXIH  OF  LEUCOCYTES  261 

local  and  general  increase  in  eosinophiles,  and  we  may  even  have 
local  mast-cell  leucocj'tosis.^^ 

Tissue  cells  were  found  by  Adler  to  migrate  far  into  blocks  of  elder 
pith,  apparently  ratlier  later  than  the  leucoc^^tes.  As  they  showed 
changes  of  form  indicating  ameboid  motions  he  considers  their  migra- 
tion to  be  an  active  process.  The  existence  of  the  polymorphonuclear 
forms  in  the  pith  seems  to  be  very  transient. 

The  position  taken  by  the  yonng  blood-vessels  and  cells  in  granula- 
tion tissue,  at  right  angles  to  the  surface,  possibly  also  depends  on 
chemotaxis  determining  the  direction  in  which  the  new  cells  shall  pro- 
liferate. 

Thermotaxis  of  Leucocytes. — Heat  seems  to  affect  leucocytes 
much  as  it  does  ameba*,  moderate  temperatures  being  positively  ther- 
motactic.  ^Mendelssohn  ^-  states  that  the  thermotaxis  is  most  pro- 
nounced at  a  temperature  of  36°-39°  C.  (97°-102°  F.),  but  is  still 
marked  as  low  as  20°  C.  Temperatures  higher  than  39°  C.  (102°  F.) 
do  not  seem  to  attract  them.  Wlassow  and  Sepp  ^^  state  that  motility 
of  leucocytes  is  increased  by  warming  to  40°  C,  and  that  temperature 
of  ■i2°-46°  C.  causes  the  movements  to  become  very  irregular,  with 
feeble  power  of  contraction.  Lymphocytes  are  not  motile  at  ordinary 
temperature,  but  at  44°  they  begin  to  move,  and  once  motile,  they 
continue  their  motion  when  cooled  as  low  as  35° ;  this  motility  is  con- 
sidered to  be  entirely  abnormal  and  only  the  result  of  degenerative 
changes.  If  mixtures  of  leucocytes  and  bacteria  sensitized  with  op- 
sonins are  kept  at  low  temperature,  the  bacteria  become  attached  to 
the  surface  of  the  leucocytes,  not  being  ingested  until  the  mixture  is 
warmed.'^*  This  indicates  that  two  separate  processes  are  involved  in 
phagoc.ytosis. 

Temperature  probably  plays  but  a  minor  part  in  attracting  leuco- 
cytes in  pathological  processes,  however.  The  local  heat  of  an  inflamed 
area  is  due  chiefly  to  the  accumulation  of  blood  in  the  part,  and  would 
not  influence  the  leucocytes  to  migrate  from  the  still  w^armer  blood 
into  the  tissues.  By  increasing  motility,  however,  the  temperature  of 
fever  may  favor  migration  and  phagocytosis,  and  local  application  of 
heat  to  inflamed  areas  may  induce  local  leucocytic  accumulation.  In 
bums  the  duration  of  the  period  of  excessive  temperature  is  usually 
too  brief  to  account  for  the  attraction  of  leucocytes  that  results ;  this 
accumulation  is  undoubtedly  due  to  the  products  of  the  resulting  cell 
degenerations. 

31  See  Milchener,  Zeit.  klin.  Med.,   1809    (37).   194;   Massaglia,   Cent.   f.  Path. 
1910    (21),  534. 

32  Rousskv  Vratch,  1903. 
33Vircho\V's  Archiv,  1904   (176),  185. 

34  Ledingham,  Proc.  Roval  Soc,  1908  (80).  188;  Sawtchenko,  Arch.  sci.  biol.. 
1910  (15),  145. 


262  INFLAMMATWX,  RFJJESERATlOy .  GROWTH 

The  influence  of  light,  mechanical  stimulation,  and  gravity  upon 
leucocytes  seems  not  to  have  been  studied.  The  phagocytosis  of  insol- 
uble non-nutritive  particles  has  been  ascribed  to  tactile  stimulation, 
but  the  details  of  the  operation  of  such  stimuli  are  unknown,  and  the 
entire  question  of  tactile  stimulation  is  unsettled.  In  experiments 
with  elder  pith  it  has  been  observed  that  leucocytes  penetrate  to  the 
center,  even  when  the  pith  contains  only  physiological  salt  solution. 
As  Adler  remarks,  it  is  difficult  to  explain  such  migration  as  due  to 
tactile  stimuli ;  but,  on  the  other  hand,  no  other  explanation  has  been 
offered. 

PHAGOCYTOSIS  s- 

The  engulfing  of  bacteria,  cells,  tissue  products,  etc.,  by  leucocytes 
seems  to  be  but  an  extension  of  the  phenomenon  of  chemotaxis.  When 
the  substance  toward  which  the  leucocyte  is  drawn  is  small  enough, 
the  leucocyte  simply  continues  its  motion  until  it  ha.s  flowed 
entirely  about  the  particle.  Later  the  particle  becomes,  as  a  rule, 
more  or  less  altered  within  the  cell,  unless  it  is  a  perfectly  insoluble 
substance,  such  as  a  bit  of  coal-dust.  This  action  upon  the  engulfed 
object  is  undoubtedly  due  to  the  action  of  intracellular  enzymes.^*' 
Protozoa  take  their  food  into  a  specialized  digesting  vacuole  which  has 
been  shown  by  Le  Dantec  ^'  (in  Stentor,  Faraiiioeciuui,  and  some  other 
varieties)  to  contain  a  strongly  acid  fluid.  JNIiss  Greenw^ood  ^*  has  also 
demonstrated  acid  in  several  forms  of  protozoa,  which  is  formed  under 
stimulation  of  injected  particles,  whether  nutritious  or  not.  ]Mouton  ^® 
has  been  able  to  extract  from  the  bodies  of  protozoa  (rhizopods)  a 
feebl}^  proteolytic  enzyme.  This  " amihodiastase,"  as  he  calls  it,  is 
active  in  alkaline,  and  faintly  acid  media,  and  digests  colon  bacilli  that 
have  been  killed  by  heat,  but  not  living  bacilli.  This  last  fact  is 
bighly  suggestive  in  connection  with  the  important  question  of  whether 
leucocytes  engulf  and  destroy  virulent  bacteria  or  only  those  that  have 
been  previously  injured  by  the  tissue  fluid.  It  was  impossible  to  se- 
cure either  invertase  or  lipase  in  extracts  of  protozoa.  Whether  bac- 
teria are  digested  in  leucocytes  by  the  same  enzymes  that  digest  the 
leucocytes  themselves  after  they  are  killed  {i.  e.,  the  autolytic  fer- 
ments), or  by  some  specialized  enzyme,  is  not  known.  ]\Ietclinikotf, 
however,  has  noted  the  localized  production  of  acid  in  the  cytoplasm 
of  leucocytes  of  the  larva  of  Triton  taeni-atits.  The  eventual  excre- 
tion of  the  remains  of  the  bacteria  or  other  foreign  bodies  by  tlie 

35  Sco  review  by  MotsclinikofT,  Kollo  and  Wassorniaiin's  llaiulh.  d.  Patli.  ^^ik- 
roorf^anisnien,  1!)1.3  (IT),  t;").") ;  also  II.  J.  TTainl)iirm'r.  ""PIin  sikaliscli-clu'inische 
Unt('rsucluinf,'-on  iiber  Pliapocyten,"  Bergmann.  Wioshadon,  1012.  wliere  is  given 
a  fiill  account  of  the  author's  important  researches  on  the  ])riiu'iples  of  phagocytic 
behavior. 

30  See  Opie,  Jour.  Exp.  IMed.,   lOOf)    f8).  410. 

37  Ann.  d.  I'Inst.  Pasteur,   1890    (4),  776. 

38  Jour,  of  Physiol..  1804    (16).  441. 

30  C.  R.  Acad.'des  Sciences,  1901    (13.3),  244. 


PlIACOCYTOf^IS  263 

])luifjoc'ytes  is  aseribod  by  Rhuinbler  tu  changes  in  tlic  eoiuixjsitiuu  of 
the''l)artick's  through  digcstioii,  so  that  they  have  a  greater  surface 
affinity  for  the  surrounding  fluids  than  for  the  protoplasm  of  the  cell. 
Calcium  and  magnesium  salts  increase  phagocytosis  and  leucocytic 
migration/"  while  changes  in  osmotic  pressure  decrease  these  activi- 
ties, as  also  does  (luinine  even  in  dilutions  of  0.001  per  cent.  Phago- 
cytosis cannot  take  place  in  the  absence  of  electrolytes,  according  to 
Sawtchenko.'^  Fat-soluble  substances  in  general  increase  phagocyto- 
sis (Hamburger),*''^  but  cholesterol  inhibits  phagocytosis,""^  (its  ef- 
fects being  suppressed  by  lecithin)  *''  acting  apparently  by  virtue  of 
its    OH    group.     Agents    facilitating    oxidation    favor    phagocytosis 

(Arkin)."'' 

Phagocytosis  cannot  be  readily  a.scribed  to  chemotaxis,   however, 
in  the  case  of  phagocytosis  of  perfectly  insoluble,  chemically  inert  par- 
ticles, such  as  coal-dust.     The  leucocytes  seem  to  take  up  foreign  bod- 
ies without  reference   to  their  nutritive  value,    absorbing   India-ink 
granules  and  bacteria  impartially  when  they  are  injected  together, 
and  loading  themselves  so  full  of  carmine  granules  that  they  cannot 
take  up  bacteria  subsequently  injected.     It  is  possible  that  foreign 
bodies  first  become  coated  with  a" layer  of  altered  protein  which  then 
leads  to  phagocytosis,  but  there  is  no  sufficient  evidence  for  this  sur- 
mise.    Kite  and  Wherry  "«  state  that  leucocytes  take  up  carbon  parti- 
cles and  similar  substances  because  the  leucocytes  are  "sticky,"  which 
presumably  is  correct,  but  what  constitutes  the  "stickiness"  and  why 
it  varies  under  the  influence  of  serum  is  not  indicated.     Presumably 
it  represents  an  altered  viscosity,  which  is  known  to  be  increased  by 
increased  acid  content  such  as  might  be  produced  by  local  asphyxia."*'' 
The  nature  of  mechanical  stimulation  of  cells  is  explained  by  Oster- 
hout"^  as  a  chemical  reaction  to  rupture  of  semipermeable  cellular 
surfaces,  and  there  is  evidence  from  plant  cells  supporting  this  hy- 
pothesis, but  its  applicability  to  animal  cells  has  not  been  investigated. 
The  experiments  of  Schaeffer  "^  seem  to  show  that  ameba^  exhibit 
positive  chemotaxis  towards  such  insoluble  substances  as  carbon  parti- 
cles and  glass  fragments,  even  at  a  distance,  although  the  mechanism 
is   unexplained.     Similar   investigations   have   not    been   made   with 
leucocvtes. 

Not  only  leucocytes  but  tissue  cells  are  capable  of  moving  and  per- 

40  Hamburger,   Biochem.   Zeit.,   1010    (2G),  66;   Eggers,   .Tour.  Infect.   Diseases, 

^'^I'^Ardi".  S'biol.   St.  Petersburg,    mil    (10),   161 ;  .Ifl^    HJ)  •   128. 

4ia  Ilanil.urger  and  de  Haan.  Arcli.  Anat.  und  Physiol._.  1913,  Phys.  AM.,  p.   ,,. 

41b  Dewev  and  Xu/Aim,  .Tour.  Tnfect.  Dis  ,  1014   (1.5),  472. 

4ieStuber.  Biochem.  Zeit.,  1013   (51),  211;    1014    (53),  493. 

4id  Jour.  Infect.  Dis..  1013    (13),  418. 

4ieJour.  Infect.  Dis.,  1915   (16).  109. 

4ifSee  Woolley,  Jour.  Amer.  Med.  Assoc,  1914   (63),  22/9. 

4igProc.  Natl.'  Acad.  Sci.,  1916   (2),  237. 

4ihBiol.  Bull.,  1916   (31),  303. 


264  IXFLAMMATIUX,  JiEGEXERATWX,  GlWWTH 

formiiig-  phagocj'tosis  when  properW  stimulated,  and  apparently  all  or 
nearly  all  lixed  cells  may  act  as  phagocytes  under  some  conditions. 
Their  power  of  independent  movement  is  much  less  than  their  phago- 
cytic power.  Endothelial  cells  are  particularly  active  in  phagocyto- 
sis, as  also  are  the  new  mesodermal  cells  produced  in  inflammation. 
Apparently  they  obey  the  same  laws  as  the  leucocytes,  and  not  only 
take  up  bacteria,  but  also  fragments  of  cells  and  tissues,  red  corpus- 
cles, and  even  intact  leucocytes  and  other  cells.  Brodie  •*-  considers 
that  phagocj'tosis  by  endothelial  cells  in  lymph-glands  is  the  natural 
end  of  the  leucocytes,  and  red  corpuscles  seem  to  have  a  similar 
fate. 

Phagocytosis  is  usually  accomplished  solely  by  the  cytoplasm  of  the 
cells,  the  nuclei  maintaining  a  passive  role;  but,  according  to  Detre 
and  Selli,^^  the  phagocytosis  of  particles  of  lecithin  is  accomplished  by 
the  nuclei,  which  seem  to  have  a  specific  affinity  for  this  substance. 

Giant-cell  formation  may  also  be  considered  as  the  result  of  chemo- 
taxis,  the  cells  moving  toward  the  attracting  particle,  and  when  the 
particle  is  larger  than  the  cells  they  spread  out  upon  its  surface,  their 
cytoplasm  flowing  together  because  of  altered  surface  tension.  The 
peripheral  disposition  of  the  nuclei  probably  depends  on  the  fact 
that  in  ameboid  motion  the  nucleus  of  the  cell  plays  an  entirely  passive 
role,  being  dragged  along  by  the  cytoplasm,  and  hence  it  is  located 
most  remotely  from  the  attracting  particle.  Digestion  of  materials 
taken  into  a  giant-cell  seems  to  go  on  as  in  the  individual  cells  that 
compose  it.^* 

Influence  of  the  Serum  on  Phagocytosis  (Opsonins). — Phago- 
cytosis of  bacteria  by  leucocytes  seems  not  to  be  merely  a  reaction  be- 
tween the  leucocytes  and  the  bacteria.  Wright  and  Douglas  have  dem- 
onstrated that  certain  substances  in  the  blood-serum  are  necessary  to 
I)repare  the  bacteria  for  phagocytosis,  these  substances  being  termed 
by  them  ''opsonins."  If  leucocytes  are  washed  free  from  serum  with 
salt  solution  and  let  stand  in  a  test-tube  with  such  bacteria  as  Strep- 
tococcus pyogenes,  Staphylococcus  pyogenes,  B.  typhosus,  B.  coli,  B. 
tuherculosis,  and  various  other  organisms,  no  phagocytosis  occurs.  If, 
however,  some  serum  from  a  normal  or  an  immunized  animal  is  added 
to  the  mixture,  active  phagocytosis  soon  takes  place.  The  action  of 
opsonins  is  also  involved  in  phagocytosis  by  endothelium.^'  The  char- 
acter and  properties  of  the  opsonins  are  further  considered  among  the 
reactions  of  immunity  (Chapter  vii). 

Results  of  Phagocytosis. — .After  phagocytosis  has  been  accom- 
plished, the  fate  of  the  cir'nlfed  object  dei)ends  ujion  its  nature.  Tf 
digestible  by  the  intracelhdar  enzymes  it  is  soon  destroyed,   but   in 

42  Jour,  of  Anat.  and  Phvsiol.,  1901    (.•?;-)),  142. 
4.^T{«rl.  klin.  \\ofh.,  ]nn.5"(42).  940. 

44  Spe  Fabcr.,  Jour,  of  Path,  and  Pact.,  1S9.3    (1).  .149. 
4'' Priscoe,  Jour.  Path,  and  Pact.,  1907    (12),  fit!. 


PHAGOCYTOSIS  265 

ihe  case  of  engulfed  living  cells,  it  seems  probable  tbat  tliey  must  be 
first  killed — they  form  no  exception  to  the  rule  that  living  protoplasm 
t-annot  be  digested.  This  brings  forward  the  (juestion  of  so  much 
importance  in  the  problems  of  innnunity :  Do  living  bacteria  enter 
phagocytes,  or  are  they  first  killed  by  extracellular  agencies  before 
they  can  be  taken  up  ?  At  the  present  time  it  seems  to  be  positively 
established  that  leucocytes  do  take  up  bacteria  which  are  still  viable, 
and  which  may  either  grow  inside  the  leucocyte  or  may  be  destroyed 
by  intracellular  processes.^"  On  the  other  hand,  leucocytes  do  not 
take  up  extremel}"  viiiilent  bacteria,  and  hence  the  question  as  to  the 
relative  importance  played  by  the  leucocyte  and  b}'  the  body  fluids  is 
still  undetermined.  It  is  probable  that  phagocytosis  by  fixed  tissue- 
cells  is  of  much  less  importance  in  checking  bacterial  growth  than  is 
phagocytosis  by  leucocytes.  Thus  Ruediger's  experiments  showed 
that  emulsions  of  organs,  with  the  exception  of  bone-marrow,  do  not 
destroy  streptococci  which  are  readily  destroyed  by  leucocytes.  How- 
ever, the  phagocytic  activity  of  certain  endothelial  cells,  especially  in 
lymph  sinuses  and  the  Kupffer  cells  of  the  liver,  is  so  great  that  these 
cells  may  equal  or  surjDass  the  leucocytes  in  bactericidal  power. 
Leucocytes  do  not  seem  to  bind  bacterial  toxins.^" 

Indigestible  substances  may  remain  in  cells,  particularly  in  fixed 
tissue  cells,  for  very  long  periods,  if  the  substances  are  chemically  in- 
ert. The  leucocytes  seem  to  transfer  the  indigestible  particles  which 
they  have  engulfed  to  other  tissues,  particularly  to  the  lymph-glands ; 
this  is  probably  accomplished  by  phagocytosis  of  the  laden  leuco- 
cytes by  the  macrophages  of  the  lymph  sinuses,  but  how  the  insoluble 
particles  are  later  transferred  to  the  gland  stroma  or  perihT^nphangial 
tissues,  where  they  are  chiefly  found  in  such  conditions  as  anthracosis, 
etc.,  is  quite  unknown. 

Leucocytes  contain  substances  which  are  strongly'  bactericidal,  in- 
dependent of  the  action  of  the  blood  serum,  and  which  have  been 
called  cndolysins;  *^  they  are  resistant  to  65°  or  even  higher,  and  seem 
to  be  bound  rather  firmly  to  the  protoplasm  of  the  leucocytes,  for  they 
resist  extraction  except  by  vigorous  methods;  they  have  a  complex 
structure  like  the  amboceptor-complement  bacteriolysins  of  the  serum,, 
and  are  not  specific  (Weil).*'*  They  do  not  pass  through  porcelain 
filters  readily,  are  precipitated  by  saturation  with  ammonium  sul- 
phate, and  resemble  the  enzymes  in  many  respejcts.^°  It  is  probable 
that  the  endolysins  act  upon  bacteria  that  have  been  phagocyted,  and 
perhaps  also  upon  free  bacteria  when  liberated  in  suppuration  through 

46  See  Riiediger,  Jour.  Amer.  Med.  Assoc,  100.5    (44).  108. 

•*'■  Fetter sson,  Zeit.  Imnuinitiit..  1011  (8),  408.  Kohzarenko,  however,  states 
that  horse  leucocvtes  neutralize  diphtheria  but  not  tetanus  toxin.  (Ann.  Inst. 
Pasteur,  1915   (29).  190.) 

*s  For  general  review  see  Kling,  Zeit.  Ininiunitat.,  1910   (7),  1. 

49  Arch.  f.  Hyg.,  1011   (74),  289. 

soManwaring,  Jour.  E.xp.  Med.,   1912    (16),  250. 


266  IXFLAMMATION,  REGENERATION,  GROWTH 

disiiitegratiou  of  the  leucocytes.     Lymphocytes  and  macrophages  seem 
to  be  devoid  of  this  endolysin.'^^ 

Phagocytosis  of  living  virulent  bacteria  may  not  always  be  an  un- 
mixed benefit.  Besides  the  obvious  pos.sibility  of  transporting  the 
bacteria  and  spreading  infection,  we  have  also  evidence  that  living 
bacteria  may  be  protected  through  phagocytosis,  against  the  action  of 
bactericidal  substances  in  the  blood  and  tissues  (Rous  and  Jones)  ."^ 

THEOEIES  OF  CHEMOTAXIS  AND  PHAGOCYTOSIS 

On  the  assumption  that  leucocytes  obey  the  same  laws  in  their  mo- 
tions as  do  the  ameba?,  studies  of  the  latter  and  of  other  forms  of 
protozoa  have  furnished  most  of  the  ideas,  hypotheses,  and  theories 
of  the  forces  involved  in  leucocytic  activities.  The  structural  rela- 
tion of  the  leucocyte  to  the  ameba  is  striking,  although  by  no  means 
complete;  the  relation  of  their  activities  is  even  closer.  Each  is  a 
microscopic,  independent,  unicellular  organism,  moving  freely  in  all 
directions  by  means  of  pseudopodia  and  protoplasmic  streaming, 
taking  other  smaller  bodies  into  its  substance  and  digesting  them, 
reacting  similarly  to  like  stimuli,  and  containing  similarly  a  nucleus 
and  many  granules.  The  differentiation  of  the  protoplasm  of  the 
ameba  into  a  clear  outer  ectosarc  and  an  inner  granular  endosarc  is 
perhaps  an  important  difference,  but  so  far  as  the  two  forms  of  cells 
have  been  studied,  the  elTect  of  this  difference  in  structure  does  not 
seem  to  have  been  considered.  That  the  unicellular  protozoa,  devoid  of 
any  central  nervous  system,  and  without  any  apparent  co-ordinating 
mechanism,  seem  able  to  move  about  in  a  purposeful  way,  going  toward 
food  supplies  and  away  from  injurious  agencies,  toward  or  away  from 
light,  heat,  and  chemicals,  has  long  attracted  the  interest  of  physi- 
ologists, particularly  as  in  these  single-celled  organisms  w^e  may  look 
for  the  simplest  conditions  of  existence  and  the  most  elementary  life 
processes.  It  seems  absurd  to  imagine  that  a  paramaecium  goes  toward 
a  dilute  acid  because  it  "likes  it,"  that  an  ameba  rejects  a  piece  of 
glass  because  it  "does  not  taste  good,"  as  we  explain  similar  nuuii- 
festations  in  higher  forms ;  furthermore,  it  has  been  shown  by  Verworn 
that  minute  enucleated  fragments  of  protozoan  cells  react  to  stimuli 
just  as  does  the  entire  cell,  and,  therefore,  it  seems  that  the  only  possi- 
ble explanation  of  movements  in  protozoa  must  be  a  direct  reaction 
of  the  stimuhited  part  to  the  stimulus.  The  luiture  of  the  stimulus 
and  the  nature  of  the  stimulated  substance  must  determine  the  nature 
f)f  the  resulting  reaction,  and  most  of  the  observations  so  far  made 
suggest  that  these  reactions  can  be  explained  according  to  the  known 
law^s  of  tlie  physics  of  fhiids.  An  ameba,  or  a  leucocyte,  may  be  lookeil 
upon  as  a  di'oj)  of  a  colloidal  solution,  surrounded  l)y  a  delicate  sur- 

M  Sco  Schneider,  Arch.  f.  Tlvpr.,   1000    (70),  40. 
sia.Iour.  Exper.  Med.,  lOlG   (23),  GOl. 


ARTIFICIAL   IMITATIOSH  OF  AMFli'Hn   .\/0\  FMFST  267 

face  layer  which  is  more  or  less  readily  permeable  to  solvents  and  to 
substances  in  solution,  and  suspended  in  a  fluid  of  quite  different  com- 
position. 

Surface  Tension. — Such  a  drop  of  fluid  suspended  in  another  diirerent  fluid 
obevs  well  known  laws  of  i)iivsics.  Tlie  pailicles  of  each  fluid  are  all  under  the 
influence  of  a  verv  considerable  force,  called  the  cohesion  pressure,  wincli  tends 
to  draw  tliem  to^nHlier  closely.  Within  the  drop  each  particle  is  subjected  to 
this  force  alike  from  all  sides,  so  that  the  forces  neutralize  one  anotber,  and 
each  particle  is  as  free  as  if  there  were  no  cohesion  pressure.  But  the  particles 
on  the  surface  are  subjected  to  unequal  pressure,  for  that  of  the  fluid  outside 
the  drop  is  difl'erent  from  that  inside,  and  so  the  pressure  on  the  surface  particles 
is  equal  to  the  diflerence  of  the  cohesion  pressure  of  the  two  fluids:  tins  con- 
stitutes the  surface  tension.  It  is  this  tension  that  pulls  in  upon  the  surface 
continuallv,  causing  it  to  tend  always  to  reduce  the  free  surface  to  a  niinnnum, 
which  condition  exists  perfectly  in  the  sphere.  The  amount  of  cohesion  aflinity 
is  yery  diflerent  in  difl'erent  fluids,  and  therefore  some  haye  a  high  surface 
tensioii  and  some  a  low.  When  a  substance  dissohes  in  another  the  surface  ten- 
sion is  a  resultant  of  the  surface  tension  of  the  two  substances,  and  hence  the 
surface  tension  of  a  liciuid  may  be  raised  or  lowered  by  dissolying  yarious  sub- 
stances in  it. 

ARTIFICIAL  IMITATIONS  OF  AMEBOID  MOVEMENT 
Imagine  a  drop  of  fluid  suspended  in  water— let  it  be  a  drop  of 
protoplasm,  or  oil,  or  mercury;  the  drop  owes  its  tendency  to  as- 
sume a  spherical  shape  to  the  surface  tension,  which  is  pulling  the  free 
surface  toward  the  center  and  acting  with  the  same  force  on  all  sides. 
The  result  is  that  the  drop  is  surrounded  by  what  amounts  to  an 
elastic,  well-stretched  membrane,  similar  to  the  condition  of  a  thin 
rubber  bag  distended  with  fluid.  If  at  any  point  in  the  surface  the 
tension  is  lessened,  while  elsewhere  it  remains  the  same,  of  necessity 
the  wall  will  bulge  at  this  point,  the  contents  will  flow  into  the  new 
space  so  offered,  and  the  rest  of  the  wall  will  contract ;  hence  the  drop 
moves  toward  the  point  of  lowered  surface  tension.  Conversely,  if 
the  tension  is  increased  in  one  place,  the  wall  at  this  point  will  eon- 
tract  with  greater  force  than  elsewhere,  driving  the  contents  toward 
the  less  resistant  part  of  the  surface,  and  the  drop  will  move  away 
from  the  point  of  increased  tension.  The  resemblance  of  these  changes 
of  form  and  the  type  of  motion  produced,  to  ameboid  movement,  is 
apparent,  and  much  experimenting  has  been  done  to  determine  how 
far  the  processes  of  motion  as  shown  by  ameba?  and  leucocytes  can 
be  reproduced  by  fluid  drops  under  various  conditions  of  experiment, 
and  to  ascertain  if  such  ameboid  movement  of  living  cells  can  be 
entirely  explained  by  the  laws  of  surface  tension. 

Gad'"'-  in  1878,  pointed  out  the  resemblance  to  ameboid  motion  of 
the  changes  in  shape  observed  in  drops  of  rancid  oils  in  weak  alkaline 
solution.  These  changes  in  shape  are  due  to  the  formation  of  soaps 
which  lower  the  surface  tension  of  the  drop  in  places,  so  that  the 
fluid  flows  toward  these  places  and  produces  pseudopodium-like  pro- 
jections. 

52DuBois  Reymond's  Arch.   f.  Physiol.,    1S7S,   p.   ISl. 


268  JXFLAMMATIOX,  REGENERATION,  OROWTH 

G.  Quincke  "^  later  ascribed  the  contractions  and  other  movements 
of  ameba?  to  alterations  of  the  surface  tension  of  the  living  substance 
in  relation  to  that  of  the  surrounding  medium,  believing  the  sub- 
stances responsible  for  the  alterations  to  be  albuminous  soaps. 

Biitsehli  ^*  found  that  drops  of  "foam  structure"  made  by  mixing 
rancid  oil  and  potassium  carbonate  solution  show  '"protoplasmic 
streaming"  when  placed  in  glycerol,  and  that  they  exhibit  positive 
chemotaxis  toward  soap  solution  and  other  chemicals,  the  motion  be- 
ing accompanied  by  current  formation  in  the  drops.  The  "pseudo- 
podia"  formed  by  the  drops  also  show  currents  rushing  along  their 
axes  and  returning  by  way  of  the  surface.  Heat  leads  to  increased 
activity  of  motion.  The  motions  were  ascribed  by  Biitschli  to  the 
bui*sting  of  some  of  the  superficial  globules  of  the  foam,  which  then 
spread  over  the  surface  of  the  drops,  lowering  its  surface  tension  at 
the  point  of  contact.  He  believed  that  ameboid  motion,  likewise, 
depended  upon  rupture  of  surface  globules  of  protoplasm,  for  the 
"foam  structure"  of  which  he  has  been  the  leading  advocate. 

Bernstein, ^^  basing  his  work  on  some  observations  of  Paalzow,  ob- 
served that  a  completely  inorganic  substance,  a  drop  of  quicksilver, 
could  be  made  to  imitate  ameboid  motion  under  the  influence  of 
chemical  changes.  If  a  crystal  of  potassium  dichromate  is  placed 
near  a  drop  of  quicksilver  in  a  nitric  acid  solution,  as  soon  as  the 
yellow  color  made  by  diffusion  of  the  dichromate  reaches  the  drop  the 
quicksilver  begins  to  show  motion  and  advances  toward  the  crystal. 
This  movement  is  due  to  local  oxidation  of  the  surface  mercury,  which 
lowers  the  tension  on  that  side  of  the  drop,  toward  which  the  mercury 
then  flows.  If  the  crystal  is  removed,  the  drop  follows,  often  flow- 
ing about  it  as  if  to  take  it  in,  but  soon  again  withdrawing  when  the 
acid  dissolves  away  the  oxide  formed  on  the  surface,  only  to  return 
again  later.  All  these  movements,  which  may  be  very  life-like,  are 
readily  explained  by  changes  in  surface  tension  that  take  place  under 
the  influence  of  the  bichromate  and  the  acid,  and  are  unquestionably 
referable  to  surface  phenomena. 

Artificial  Amebae. — By  far  the  most  suggestive  experiments  on 
the  simulation  of  ameboid  activity  by  non-living  substances  are  those 
of  Rhumbler  (1898)  in  his  great  work,  "  Pliysikalische  Analj'se  von 
Lebenserscheinungen  der  Zelle. "  ''^  On  tlie  assumption  that  the  living 
])rotoplasm  was  but  a  more  or  less  tenacious  fluid,  following  the 
simple  pliysical  laws  of  fluids,  especially  in  relation  to  its  surface  ten- 
sion, he  devised  a  number  of  experiments  to  determine  the  correctness 
of  these  viewg.  An  ameba  may  be  regarded  as  such  a  mass  of  viscid 
fluid,  in  a  medium  in  wliich  it  is  nearly  or  (|uite  insoluble;  it  is  also 

03  Wio<lniaiin"s  Aiinalcii.    ISSS    (.35),   oSO. 

54  "I'ldtojiliisiii."  translation  liy  ^fincliin.  T^ondon,   1804. 

55  Pfliifjer's  Arcli.,  1!)00   (80),  628. 

50  Arch.  f.  Entwic-klungsmechanik,   1808    (7),   103. 


ARTIFICIAL  AMEB/E  269 

constantly  undergoing  chemical  changes  within  itself,  and  taking  sub- 
stances from  or  secreting  tliem  into  the  surrouiuliiig  water.  To  re- 
produce partly  these  conditions  a  drop  of  clove  oil  is  placed  in  a 
mixture  of  glycerol  and  alcohol ;  the  alcohol  and  clove  oil  are  miscible, 
the  glycerol  merely  retarding  the  diffusion.^"  Such  a  drop  of  oil  will 
move  about,  changing  its  form  and  sending  out  pseudopotlia  nmch  as 
an  ameba  does.  These  movements  are  undoubtedly  due  to  changes 
in  the  surface  tension  brought  about  by  the  irregular  mixing  of  the 
alcohol  and  the  clove  oil.  The  effect  of  chemotaxis  upon  an  ameba 
can  likewise  be  imitated  with  such  an  "artificial  ameba."  If  some 
stronger  alcohol  is  carefully  introduced  into  the  fluid  near  the  drop, 
the  surface  tension  on  that  side  will  be  lowered,  and  the  drop  will 
flow  in  that  direction.  The  effect  of  chemical  changes  within  the  drop 
upon  its  motion  may  be  demonstrated  similarly  by  injecting  a  little 
alcohol  into  the  substance  of  the  drop  near  one  edge — the  drop  will 
send  out  a  pseudopodium  on  that  side,  and  perhaps  flow  along  in 
the  direction  of  the  pseudopodium.  AYe  can  imagine  that  metabolic 
changes  in  the  body  of  an  ameba  may  account  for  many  of  its  seem- 
ingly purposeless  movements  by  altering  surface  tension  in  some  part 
of  its  circumference.  Thermotaxis,  the  effect  of  heat  in  modifying  or 
impelling  ameboid  motion,  may  be  equally  well  demonstrated  in  such 
an  "artificial  ameba,"  the  drop  being  "positively  thermotactic,"  and 
flowing  rapidly  toward  a  heated  point  in  the  solution,  because  heat 
lowers  the  surface  tension. 

Even  as  highly  specialized  a  process  as  the  taking  of  food  may  be 
closely  simulated  experimentally.  Ameba'  seem  to  possess  the  faculty 
of  selecting  substances  that  are  suitable  for  their  food,  crawling  over 
particles  of  sand,  wood,  etc.,  and  rejecting  them  when  they  are  pushed 
against  or  into  the  surface  of  the  ameba,  which,  however,  readily  takes 
up  bacteria,  diatoms,  algfe,  etc.,  digests  them,  and  later  throws  out  the 
undigested  particles.  If  there  is  any  property  of  the  ameba  that  sug- 
gests voluntary  action,  it  seems  to  be  exhibited  in  the  choice  of  its 
food,  although  this  is  not  so  well  developed  a  selective  process  as 
might  be  expected,  for  amebje  will  take  up  many  harmful  objects,  and 
they  may  be  made  to  fill  themselves  so  full  of  useless  substances  that 
they  cannot  take  up  food.  However,  a  drop  of  chloroform  in  water, 
which  makes  a  good  artificial  ameba,  if  "fed"  with  various  substances, 
will  refuse  some  and  take  in  others  in  a  surprisingly  life-like  manner. 
Pieces  of  glass  or  of  wood  placed  in  contact  with  the  drop,  exert  no 
influence ;  if  pushed  into  the  substance  of  the  drop,  they  carry  the 
surface  ahead,  and  on  being  released  they  are  throw^i  out  with  some 
force.  If  a  piece  of  shellac,  paraflln,  styrax,  or  Canada  balsam  be 
brought  in  contact  with  the  surface  of  the  drop,  however,  the  drop 
flows  around  it  immediately,  and  takes  it  within  its  substance,  where 

57  The  details  of  these  experiments  are  as  given  briefly  hx  Jennings.  Jour,  of 
Applied  Microscopy,  1902    (5),  1597. 


270  J X FLA  MM ATIOX,  REGEyERATIOy,  GROWTH 

it  is  soon  dissolved.  Even  more  strikingly  like  phagocytosis  and  in- 
tracellular digestion,  however,  is  the  result  of  a  similar  experiment 
Avith  a  piece  of  glass  covered  with  shellac;  the  chloroform  "ameba" 
takes  it  up  as  readily  as  it  does  the  shellac  alone,  but  after  all  the  coat- 
ing is  dissolved  aAvay  the  piece  of  glass  is  then  cast  out  of  the  drop. 
The  resemblance  to  the  engulfing,  digestion,  and  excreting  of  indigesti- 
ble particles  of  bacteria,  etc.,  by  amebic,  is  so  striking  that  it  seems 
impossible  that  there  can  be  any  fundamental  differences  in  the  two 
processes.  It  will  also  be  noticed  that  the  drop  takes  in  only  what 
it  can  dissolve  and  rejects  what  it  cannot. 

One  of  the  most  remarkable  actions  of  the  ameba^,  which  seems  al- 
most certainly  the  result  of  voluntary  action,  is  this:  Oftentimes  in 
feeding,  an  ameba  gets  hold  of  a  suitable  material  which  is  in  the 
form  of  a  long  thread,  much  too  long  for  the  ameba  to  surround.  It 
then  proceeds  to  coil  up  the  thread  within  its  body,  by  stretching  a 
slight  distance  along  the  thread,  bending  over,  and  forming  a  bend  in 
the  thread,  and  by  repeating  the  process  it  crowds  the  thread  into  a 
neat  coil  within  its  body,  where  it  can  be  digested.  The  process  is 
done  so  systematically  and  with  such  evident  adoption  of  the  means 
at  hand  to  the  desired  end,  that  it  seems  as  if  it  must  be  an  adaptation 
of  the  ameba  to  circumstances,  the  result  of  long  experience  or  of 
heredity.  That  an  artificial  ameba  can  perform  the  same  maneuvers 
seems  hardly  credible,  but  it  is  readily  done  with  almost  no  difference 
in  detail.  If  the  chloroform  drop  is  given  a  long  fine  thread  of  shel- 
lac, it  proceeds  to  bend  the  thread  in  the  middle,  and  to  send  pseudo- 
podia  out  along  the  thread  to  pull  it  into  the  drop,  coiling  it  up  inside 
as  the  chloroform  softens  the  substance  of  the  thread,  until  it  is  all 
contained  within  the  drop,  provided,  of  course,  that  it  is  not  too  long 
(a  thread  six  times  as  long  as  the  chloroform  drop  may  be  taken  in 
completely).  The  bending  and  coiling  of  the  thread  in  this  experi- 
ment is  entirely  in  accord  with  the  known  laws  and  phenomena  of 
surface  tension. 

Fully  as  striking  an  ameboid  action  as  the  coiling  up  of  a  thread 
too  long  to  be  taken  in,  is  the  building,  by  some  of  the  protozoa  closely 
related  to  the  ameba  (Difflugia)  of  a  shell  which  the  animal  seems 
to  form  by  cementing  together  grains  of  sand,  or  diatom  shells,  or 
other  suitable  particles.  Tlie  particles  are  united  so  closely  and  fitted 
together  so  well  that  tliey  are  almost  perfectly  free  from  crevices. 
Even  this  process  is  accurately  imitated  in  Rhumbler's  experiments. 
If  a  drop  of  oil  is  mixed  with  fine  grains  of  quartz  sand,  and  droi)ped 
into  70  per  cent,  alcohol,  the  grains  are  thrown  out  to  the  surface, 
where  they  adhere  to  the  surface  of  the  drop  and  to  one  anotlier 
exactly  as  do  the  particles  in  a  difflugia  shell.  So  well  fitted  are  tlie 
particles  that  the  artificial  shell  may  remain  intact  for  months,  and 
resemble  the  natural  slicll  indistinguishably. 

Furthermore,  the   phenomenon  of  cell   (li\ision  can  be  imitated  to 


AMEBOID    MOT/OX  OF  LEUCOCYTES  271 

some  extent  by  oil  di-oplcts.  liiitsdili  considered  that  tlie  cleavajic 
furrow  of  dividino:  cells  represented  an  area  of  grreater  surface  ten- 
sion, and  iMcClendon  imitated  cell  division  as  follows:  He  suspended 
a  drop  of  rancid  oil  and  chloroform  between  water  and  salt  solution, 
and  allowed  sodium  liydi'ate  to  floAv  from  pipettes  against  two  oppo- 
site points  in  the  droplet,  whereon  the  surface  tension  was  lowered 
and  the  drop  bulged  at  these  points,  the  band  of  higher  surface  tension 
constricting  the  drop  between  these  two  points.  Burrows  states  that 
the  changes  seen  in  cells  dividing  beneath  the  microscope  correspond 
well  to  tliese  experimental  observations.^'*^ 

RELATION    OF    THE    ABOVE    EXPERIMENTS    TO    THE    PHENOMENA    EX- 
HIBITED BY  LEUCOCYTES  IN  INFLAMMATION 

The  experiments  cited  indicate  strongly,  to  say  the  least,  that  amebse, 
and  presumably  leucocytes,  react  to  stimuli  of  various  kinds,  chietly 
through  the  effect  of  these  stimuli  upon  surface  tension.  The  stimuli 
may  come  from  within  the  cell,  being  in  this  case  the  result  of  changes 
in  composition  brought  about  by  metabolic  processes;  such  chemical 
products  alter  the  tension  of  the  surface  nearest  their  point  of  origin, 
causing  what  appears  to  be  spontaneous  motion.  Stimuli  acting  from 
without  may  be  chemical,  thermal,  electrical,  or  mechanical,  but  in 
any  event  they  act  as  stimuli  to  motion  through  their  effect  upon  sur- 
face tension ;  if  they  decrease  the  surface  tension  the  cell  goes  toward 
them ;  if  they  increase  the  tension,  the  cell  moves  away.^''*'  The  be- 
havior of  leucocytes  in  inflammation  may  be  explained  on  these  purely 
physical  grounds  very  satisfactorily,  as  follows: 

At  the  point  of  cell  injury  or  of  infection,  substances  are  produced 
that  exert  positive  chemotaxis.  as  can  be  shown  by  experiments  both 
outside  and  inside  the  body ;  these  substances  are  chemotaetic  because 
they  influence  the  surface  tension  of  the  leucocytes,  and  since  with 
most  if  not  all  the  products  of  cell  disintegration  the  effect  is  to  lower 
surface  tension,  the  chemotaetic  effect  is  positive.  As  the  chemotaetic ' 
substances  are  produced,  they  diffuse  through  the  tissues  until  they 
reach  the  walls  of  a  capillary,  through  which  they  begin  to  pass,  pre- 
sumably most  rapidly  through  the  thinnest  parts  of  the  wall,  the 
'"stomata"  and  intercellular  substance.  The  leucocytes  passing  along 
in  the  bore  of  the  capillary  will  be  touched  by  the  chemotaetic  sub- 
stances most  on  the  side  from  which  the  substances  diffuse ;  the  sur- 
face tension  will  be  lowered  on  this  side,  causing  the  formation  of 
pseudopodia  and  motion  in  this  direction.  When  the  leucocytes  come 
in  contact  with  the  wall,  their  surfaces,  because  saturated  with  the 
chemotaetic  substances,  will  have  a  tension  much  the  same  as  that 
of  the  cells  of  the  capillar^'  wall,  which  are  likewise  saturated  with  the 

57a  See  Trans.  Cono;ress  Amef.  Phys..   1913    (9),  77. 

57b  OH-ions  decrease,  Il-ions  increase  the  surface  tension  of  leucocytes 
(Schwyzer,  Biochem.  Zeit.,  1014  (60),  306.  447,  454),  which  may  explain  the  "fact 
that  lactic  and  other  acids  exhibit  negative  chemotaxis. 


272  INFLAMMATION,  REGENERATION,  GROWTH 

same  substances,  and  the  two  surfaces  will  tend  to  cling  to  one  another ; 
explaining  the  phenomenon  of  adhesion  of  leucocytes  to  the  capillary 
wall,  when,  according  to  the  usual  description,  ''the  leucocytes  be- 
have as  if  either  they  or  the  capillary  wall  had  become  sticky. ' '  ^®  Sur- 
face tension  of  the  leucocytes  will  be  least  nearest  the  points  where  the 
most  chemotactic  substances  are  entering  the  capillary,  namely,  the 
stomata;  hence  the  pseudopodia  will  form  in  this  direction  and  flow 
through  the  openings,  the  rest  of  the  cytoplasm  flowing  after  and 
dragging  the  nucleus  along  in  an  apparently  passive  manner.  Since 
it  is  the  cytoplasm  that  seems  to  be  chiefly  affected  in  these  processes, 
the  nucleus  appearing  to  be  rendered  inert  by  its  relatively  dense  and 
fixed  structure,  the  leucocytes  with  most  cytoplasm  are  most  active  in 
emigration,  while  those  with  the  least,  the  lymphocytes,  are  affected 
relatively  little  or  not  at  all. 

Once  through  the  vessel  wall,  the  motion  continues  in  the  same 
manner,  toward  the  side  from  which  the  chemotactic  matter  comes, 
just  as  the  mercury  drop  flows  toward  the  crystal  of  potassium  dichro- 
mate,  or  the  drop  of  oil  flows  toward  the  alcohol.  If  the  leucocyte 
meets  a  substance  that  lowers  its  surface  tension  sufficiently,  it  will 
flow  around  the  object  and  enclose  it,  just  as  the  chloroform  drop 
flows  about  the  piece  of  shellac  or  balsam;  this  constitutes  phago- 
cytosis. The  motion  of  the  leucocyte  will  continue  in  a  forward  di- 
rection until  one  of  several  possible  things  happens :  (a)  The  leucocyte 
may  reach  a  point  where  the  chemotactic  substances  are  so  thoroughly 
diffused  that  the  effects  on  its  surface  are  the  same  on  all  sides; 
there  will  then  be  no  tendency  to  move  in  any  direction,  (h)  It  may 
reach  a  material  that  exerts  a  marked  positive  influence  upon  it, 
causing  much  lowering  of  the  surface  tension,  but  which  is  so  large 
that  the  cytoplasm  flowing  along  its  surface  cannot  surround  it: 
other  leucocytes  will  experience  the  same  change,  their  cytoplasm  will 
fuse  together  because  of  the  equal  lowering  of  their  surface  tension, 
and  soon  we  get  a  mass  of  leucocytes  with  fused  cytoplasm  surround- 
ing the  object,  forming  a  "foreign  body  giant-cell."  (c)  The  leuco- 
cyte may  reach  a  place  where  the  concentration  of  the  chemicals  is  so 
great  that  chemical  changes  are  produced  in  its  cytoplasm.  If  these 
changes  are  of  a  coagulative  nature,  the  surface  of  the  cell  will  be 
stiffened  so  that  it  cannot  migrate  further;  if  of  a  solvent  nature,  the 
leucocyte  is  destroyed,  (d)  It  may  reach  the  margin  of  an  area  where 
the  preceding  leucocytes  have  become  coagulated  or  otherwise  rendered 
immobile,  so  that  they  block  its  path,  while  it  is  held  fixed  by  the  at- 
traction on  this  side,  (c  and  d  explain  the  formation  of  solid  leu- 
cocytic  walls  about  areas  of  inflammation,  and  the  frequent  absence 

«8  Kreibich  (Arch.  f.  Dermatol.,  1012  (114).  585)  describes  as  chemical  chanpes 
in  the  vessel  walls  fhirinjr  the  early  stages  of  inflammation,  a  difTuse  siulanonhile 
ehanpe  throntrhoiit  the  endothelial  cells,  in  the  form  of  fine,  dust -like  particles. 
Probably  this  chancre  dejjcnds  .simply  on  an  atrurefratio!!  of  the  iiitrat'cllnlar 
lipoids. 


BEHAVIOR  OF  TISSUE-CELLS  A\D  FOUMATIOX  OF  GLiNT-CELLS       273 

of  k'lR'ocytcs  within  tlu'  central  necrotic  areas.)  (e)  The  formation 
of  cheuiotactic  .substances  may  cease  because  the  substance  causing?  the 
iutlannuation  has  been  used  up,  or  because  the  bacteria  have  been 
destroyed,  or  from  an}^  of  the  causes  that  terminate  inflammation. 
Those  leucocytes  still  advancinor  will  reach  a  point  wh(!re  there  is  as 
much  chemotactic  substance  behind  as  in  front — they  will  then  stop 
advancing.'"  As  the  fluids  exuded  in  the  central  portion  continue  to 
dilute  the  chemotactic  substances  and  wash  them  out,  there  will  soon 
be  less  chemotactic  substance  in  the  center  of  the  inflamed  area  than 
there  is  farther  out,  hence  the  leucocytes  M'ill  move  away  from  the 
center  toward  the  periphery,  following  the  chemotactic  substances  back 
into  the  blood-vessel  and  the  lymph-stream.  These  are  the  conditions 
that  exist  at  the  close  of  the  inflammatory  process,  which  results  in  the 
dispersion  of  the  leucocytes. 

General  leucocytosis  can  be  explained  equally  well  on  the  same 
grounds.  Chemotactic  substances  from  the  area  of  inflammation  enter 
the  blood-stream,  and  so,  in  a  very  dilute  form,  pass  through  the  bone- 
marrow.  The  cliemotaxis  in  the  blood  will  be  greater  than  that  of 
the  marrow,  and  the  leucocytes  will  move  toward  and  into  the  blood. 
As  long  as  the  blood  contains  more  chemotactic  substances  than  the 
marrow,  leucocytosis  will  increase,  to  stop  when  the  amount  in  blood 
and  marrow  is  alike  or  when  there  is  less  in  the  blood  than  in  the 
marrow. 

Behavior  of  Tissue=cells  and  Formation  of  Qiant=cells. — The 
free  cells  of  the  tissues  involved  in  inflammation  can,  of  course,  obey 
the  same  influences  as  the  leucocytes,  and  apparently  do  so  in  so  far 
as  they  are  not  cheeked  by  structural  impediments  to  flowing  motion ; 
1.  e.,  the  more  closely  a  cell  is  related  to  a  single  drop  of  fluid  pro- 
toplasm, the  more  closely  does  it  resemble  in  the  simplicity  of  its 
reactions  the  /'artificial  ameba."  An  illustration  of  the  cliemotaxis 
of  epithelial  cells  is  furnished  by  B.  Fischer,""  who  found  that  stained 
fats  cause  growth  and  migration  of  epithelial  cells  in  the  direction  of 
the  fat.  Cells  with  much  cytoplasm  are  best  fitted  to  move  freely,  as 
a  rule,  and  hence  we  see  chiefly  the  large  endothelial  cells  of  the  lymph 
sinuses  and  the  serous  cavities,  and  the  large  hyaline  and  granular 
••ells  of  the  blood  acting  as  phagocytes,  for  phagocytosis  is  no  difi'erent 
from  ameboid  motion  which  continues  about  a  particle  until  it  is  sur- 
rounded; likewise  we  see  the  "epithelioid"  and  large  endothelial  cells 
with  their  abundant  cytoplasm  fusing  together  to  foinn  giant-cells. 
(Note  that  such  giant-cells  are  formed  particularly  in  conditions  in 
which  the  epithelioid  cell  is  more  abundant  than  is  the  leucocyte, 
e.  g.,  tuberculosis  and  other  chronic  inflammations.     The  cells  that 

59  The  phagocytic  action  of  leucocytes  in  vitro  is  decreased  bv  substances  tliat 
lower  the  surface  tension,  e.  g.  chloroform  (Hamburger,  K.  Akad.  Wetensch.,  1911 
(XIII  (2)),  802).  Ethor-solu])le  substances  from  bacteria  have  no  effect  on 
phagocytosis    (IMiiller,  Zeit.  Immunitiit.,   1908    (I),  61). 

GoMiinch.  med.  Woch.,  1906   (53),  2041. 
18 


274  IXFLAMMATIOX,  liEaEyEh'ATIOX,  GROWTH 

fuse  about  an  infected  catgut  ligature  are  the  leucocytes,  for  they  are 
most  abundant  in  such  a  place.)  A  good  illustration,  also,  is  the 
giant-cell  formed  by  fusing  of  leucocytes  about  blastomyces  in  minute 
abscesses  in  the  epithelium  in  blastomycetic  dermatitis;  the  epithelial 
cells  cannot  flow  or  coalesce  well  because  of  their  abundance  of  stitf 
keratin  and  their  specialized  cell-wall,  and  hence  do  not  participate; 
the  leucocytes  are  individually  too  small  to  surround  the  fungus  cells, 
and  hence  they  flow  about  them  in  the  abscess  exactly  as  they  will  do 
experimentally  in  a  test-tube  or  in  a  guinea-pig's  abdomen  (Hektoen). 
The  method  of  growing  tissues  in  vitro  permits  of  observation  of  the 
process  of  giant-cell  formation,  and  establishes  that,  for  foreign  body 
giant-cells  at  least,  they  are  formed  by  fusion  of  wandering  cells 
(Lambert).*'^  The  formation  of  giant-cells  is,  on  this  ground,  but  an 
amplifieation  of  ameboid  movement  and  phagocytosis.  The  fusing  of 
the  individual  cells  is  due  to  the  lowering  of  their  surface-tension  by 
the  materials  diffusing  from  the  body  which  is  to  be  absorbed,  until 
the  surface  of  each  cell  becomes  alike,  when  the  surface  tension  at 
the  point  where  each  cell  is  in  contact  becomes  zero  and  the  cytoplasm 
runs  together. 

Objections  to  the  above  Hypothesis. — Phj^sieal  explanations  of 
ameboid  movement  seem  to  fit  very  perfectly  the  known  facts  concern- 
ing the  actions  of  leucocytes.  There  arise  but  a  few  difficulties  in  ap- 
plying these  laws  to  leucocytic  action;  one  is  the  phagocytosis  of 
chemically  inert  bodies,  such  as  coal  particles,  tattooing  materials, 
stone  dust,  etc.  We  know  that  amebae  also  may  take  up  such  inert 
materials,  although  they  generally  refuse  them,  and  it  is  believed  that 
the  particles  exert  some  local  injury  to  the  cell  wall  that  leads  to  an 
alteration  in  its  tension.  Amebee  seem  also  sometimes  to  excrete  a 
sticky  substance  over  their  surfaces  or  over  the  foreign  matter  that  is 
to  be  engulfed,  which  excretion  seems  to  be  the  result  of  surface 
stimulation.  Possibly "  leucocytes  do  the  same.  AYe  must  bear  in 
mind,  however,  that  the  protoplasmic  cells  have  much  greater  possi- 
bilities for  action  than  the  "artificial  ameba,"  since  within  the  pro- 
toplasm countless  chemical  changes  are  going  on  which  must  cause 
continual  alteration  in  surface  tension  ;  it  is  (juite  possible  that  mere 
mechanical  action  may  alter  chemical  action  at  the  point  of  contact, 
so  that  the  injuring  particle  may  become  surrounded  through  local 
liquefaction  of  the  protoplasm. 

With  the  ameba,  unfortunately,  the  explanation  of  all  its  activities 
by  purely  physical  analogies  is  apparently  not  so  successful.  Al- 
though simple  pseudopodia  may  be  produced  experimentally,  and  their 
formation  explained  readi'v  0:1  the  surface  tension  basis,  yet  we  find 
many  forms  of  pseudopodia  in  the  great  family  of  amaba\  Some  of 
them  are  brancliing,  some  are  fixed  in  extension,  some  have  a  stiff 
elastic  axis.     It  would  also  be  difficult  to  explain  cilia  as  produced  by 

61  Anatomical  Record,  1012  (G),  91. 


AMEliOllt  MOTION  OF  LEUCOCYTES  275 

changes  in  surface  tension,  yet  we  iind  in  some  protozoa  that  pseudo- 
podia  may  take  on  the  persistence  and  action  of  cilia,  and  that  cilia 
may  seem  to  chanfje  into  pseudopodia.  Jennings  has  made  a  most 
extended  stndy  of  the  relations  of  tlie  "liehavior  of  Lower  Organ- 
isms""- to  the  physical  theories  of  ameboid  motion,  and  is  unable 
to  corroborate  the  claim  that  the  processes  that  go  on  in  "artificial 
amebffi"  exactly  reproduce  those  of  living  amebaj,  or  to  accept  the 
statement  that  living  ])rotoplasm  behaves  exactly  as  any  similar  drop 
of  fluid  would  under  the  same  conditions.  He  states  tliat  the  currents 
set  up  in  artificial  amebfe  by  changes  in  surface  tension  are  not  the 
same  as  those  in  living  amebae,  contrary  to  Rhumblei*  and  to  Biitschli. 
The  movement  of  ameba,  he  maintains,  is  not  due  to  the  flowing  of 
the  contents  of  tlie  cell  in  a  central,  axial  current  out  into  the  pseudo- 
podium  and  back  on  the  sides,  as  occurs  in  the  artificial  ameba;  but 
rather  to  a  rolling  forward  of  the  upper  surface  over  the  anterior 
edge  to  the  lower  surface,  where  it  becomes  fixed  to  the  surface  on 
which  the  ameba  is  crawling.  Tlie  part  played  by  surface  tension, 
he  claims,  is  in  the  case  of  ameba?  a  very  subordinate  one,  and  it  is 
not  sufficient  to  explain  the  movements  of  the  living  cell. 

However  the  discussion  concerning  the  amebse  may  turn,  it  must 
be  appreciated  that  there  are  some  important  diff'erences  between  even 
the  ameba  and  the  leucocyte.  The  latter  has  by  far  the  simpler 
organization,  and  approaches  in  structure,  and  presumably,  therefore, 
also  in  response  to  stimuli,  more  closely  to  the  simple  drop  of  colloid 
matter.  It  has  no  pulsating  vacuoles,  no  specialized  pseudopodia, 
never  forms  shells  or  coverings,  and  does  not  conjugate  as  do  the 
amebffi.  The  extenial  surface  of  the  leucocyte  is  much  simpler,  an 
important  fact  in  connection  wdth  surface  tension  etfects,  for  in  the 
leucocyte  the  surface  seems  to  be  practically  undifferentiated,  naked 
protoplasm;  whereas  in  amebae  it  is  formed  of  a  well-differentiated 
"ectosarc, "  which  has  marked  motile  powers,  being  able  to  contract 
sufficiently  to  cut  an  injured  ameba  completely  in  two.  At  the  very 
least  the  surface  tension  explanation  of  leucocytic  action  agrees  per- 
fectlji  with  most  of  the  ohserred  actions  of  leucocjjtes,  and  it  is  the 
only  reasonable  theory  offered.  There  seems  to  be  no  middle  ground 
between  such  a  physical  theory  and  a  metaphysical  theory  which 
would  endow  a  single  cell,  without  organs  or  nervous  system,  with  the 
reasoning  powers  of  highly  developed  animals,  a  position  incompati- 
ble with  the  entire  evidence  of  experience. 

62  Publication  No.  16,  Carnegie  Institute,  Washington,  1904;  also  see  American 
Naturalist,  1904   (38),  625. 


276  IXFLAMMATION,  REGENERATION,  GROWTH 


SUPPURATION  «3 

For  the  formation  of  pus  two  couditions  are  necessary :  ( 1 )  the  ac- 
cumulation  of  leucocytes,  and  (2)  necrosis  and  liquefaction  of  cells 
and  tissue  elements.  I\Iany  leucocytes  may  be  present  in  a  tissue 
without  suppuration ;  e.  g.,  erysipelas.  Necrosis  of  cells  with  their 
g'radual  liquefaction  and  absorption  may  also  occur  without  suppura- 
tion ;  e.  g.,  infarcts,  aseptic  liquefaction  necrosis,  etc.  Hence  for  sup- 
puration to  occur  there  must  be  produced  substances  with  positive 
ehemotaxis,  to  cause  accumulation  of  leucocytes,  for  if  a  necrotic  area 
is  devoid  of  leucocytes,  it  does  not  suppurate:  e.  g.,  caseous  tubercles. 
Secondly,  necrosis  must  occur,  for  digestion  and  liquefaction  of  living 
cells  and  tissues  does  not  take  place.  Only  substances  meeting  these 
requirements — i.  e.,  causing  positive  ehemotaxis  and  cell  necrosis — 
will  cause  suppuration.  Therefore,  although  bacterial  infection  is 
the  usual  cause  of  suppuration.*'^  it  may  be  produced  hy  many  other 
substances,  among  which  the  following  are  the  best  known :  Bacterial 
proteins,  even  from  non-pathogenic  bacteria;  oil  of  turpentine,  mer- 
cury, croton  oil,  silver  nitrate  solutions  (5  to  10  per  cent.),  and  certain 
vegetable  proteins  (vegetable  "caseins"). 

An  excellent  example  of  the  importance  of  leucocytes  for  suppura- 
tive softening  is  the  caseous  tubercle,  which  is  usually  free  from 
leucocytes  and  does  not  undergo  suppuration.  If  for  any  cause  leuco- 
cytes are  attracted  into  the  caseous  area,  softening  and  pus  formation 
promptly  occur.  Hence  Heile  •'■''  found  that  while  pus  from  a  "cold" 
tuberculosis  abscess  will  not  digest  fibrin  and  does  not  give  the  biuret 
reaction,  both  reactions  appear  after  a  leucocytosis  has  been  brought 
about  by  injection  of  iodoform.  It  was  formerly  considered  that  the 
softening  was  due  to  the  digestive  action  of  the  enzymes  of  the 
infecting  bacteria,  many  of  which  were  known  to  produce  digestive 
enzymes  dissolving  protein  culture-media ;  e.  g.,  Staphylococcus  pyo- 
genea.  Although  to  some  extent  these  enzymes  may  be  a  factor  in 
causing  the  softening  of  the  fixed  tissues  and  of  the  killed  leucocytes, 
their  eflPect  is  probably  insignificant  as  compared  with  the  enzymes 
liberated  by  the  leucocytes,  as  shown  by  tlie  production  of  active 
experimental  suppuration  under  aseptic  conditions  with  turpentine, 
croton  oil,  etc.®®  Suppuration  is,  therefore,  the  result  of  three  proe- 
ms inflammatory  Exudates,  their  formation  and  oomposition.  are  considered  in 
Chapter  xii. 

04  Biieliner  considers  that  liacteria  will  not  ]>rodu('e  supjiuration  unless  they 
are  hroken  down  so  that  their  pi/ogenic  proteins  are  released:  e.  (j..  anthrax 
bacilli  cause  suppuration  when  actin^r  locally,  as  in  malipnant  pustule,  hut  not 
when  they  are  causing  septicemia,  because  only  in  the  former  case  are  their 
pvopenic  proteins  liberated. 
'"•""Zcit.  klin.  Med.,  1904  (.'">-)).  508. 
""  .\j)parently  su)iy)iirat ion  mav  occur  in  herpes  70ster  vesicles  in  the  absence 
of  bacteria,  accordin<r  tn  tlie  findinfis  of  Kreibich  (Wien.  klin.  Woch..  1001  (14) 
583 ) . 


COMPOSITION  OF  PUS  277 

esses:  (1)  Necrosis  of  cells;  (2)  local  accumulatiou  of  leucocytes; 
(3)  digestion  of  the  necrotic  cells,  fibrin,  and  tissue  elements  by  en- 
zymes which  are  derived  from  three  sources,  as  follows:  (a)  the 
leucocytes;  (&)  the  infecting  bacteria  (if  such  are  present)  ;  (c)  the 
fixed  tissue-cells.  Possibly  small  quantities  of  enzymes  are  also  intro- 
duced in  the  blood  plasma,  but  these  are  probably  very  inconsiderable. 
Normal  serum,  and  probably  also  normal  cells,  contain  antibodies  for 
the  proteolytic  enzymes  of  the  leucocytes,  and  hence  neutralization  or 
destruction  of  these  antibodies  must  be  an  important  factor  in  de- 
termining the  rate  and  amount  of  suppuration.*'^ 

The  influence  of  the  antienz^nnes  is  well  shown  by  the  rabbit,  with 
serum  rich  in  antienzymes  and  leucocytes  i)oor  in  protease,  so  that 
infections  with  pus  cocci  do  not  usually  lead  to  the  formation  of  liquid 
pus  (Oine).  In  man  we  see  a  similar  relation,  in  that  exvidates  rich 
in  serum  do  not  suppurate  because  the  enzymes  are  inliibited  by  the 
senim ;  but  if  the  excess  of  serum  is  removed  suppuration  may  then 
occur.  AVith  an  excess  of  enzyme  (i.  e.,  leucocytes)  the  inhibiting 
effect  may  also  be  overcome,  and  suppuration  then  begins.  Variations 
in  the  proportion  of  leucoprotease  and  serum  antiprotease  determine, 
therefore,  the  occurrence  of  suppuration,  and  the  inflammatory  re- 
action is  seen  to  be  fundamentally  the  same  as  the  humoral  reactions 
of  immunity,  in  that  in  each  case  the  essential  process  is  the  provision 
of  proteolytic  enzymes  to  remove  foreign  or  abnormal  protein  sub- 
stances. In  inflammation  the  proteolytic  enzymes  are  brought  in  the 
leucocytes,  in  humoral  reactions  the  enzymes  are  present  free  in  the 
plasma.  The  antiproteases  may  be  of  the  nature  of  lipoids,  probably 
with  unsaturated  fatty  acids  (Jobling). 

The  proteolytic  enzymes  of  the  leucocytes  and  tissue-cells  have  been 
previously  considered  in  connection  with  the  subject  of  autolysis 
(Chap,  iii),  and  it  is  necessary  here  only  to  call  attention  to  the  fact 
that  these  enzymes  are  of  at  least  two  varieties:  (1)  Proteolytic 
enzymes  of  the  polymorphonuclear  leucocytes,  which  act  best  in  alka- 
line medium  (Opie  ^^)  ;  (2)  autolytic  enzymes  of  the  tissue-cells,  which 
act  best  in  an  acid  medium  (Hedin,  et  al.) .  The  mononuclear  leuco- 
cytes contain,  like  the  tissue-cells,  enzymes  acting  in  an  acid  medium. 
The  antienzymatic  action  of  serum  is  favored  by  an  alkaline  reaction, 
but  is  altogether  lost  in  an  acid  medium  (Opie). 

COMPOSITION  OF  PUS 

Because  of  its  method  of  production,  pus  consists  of  the  follow- 
ing substances:  (1)  The  constituents  of  the  exuded  blood  plasma; 
(2)  the  constituents  of  the  leucocytes  (and  tissue-cells)  that  exist  free 
in  the  pus;  (3)  the  products  of  digestion  of  the  proteins  of  the  leuco- 

67  See  Opie,  Jour.  Exper.  Med.,  1905  (7),  316;  1907  (9),  207;  Arch.  Int.  Med., 
1910  (5),  541. 

68  Jour.  Exper.  Med.,  1906   (8),  410. 


278  IXFLAMMATIOX,  REGENERATIOX,  GROWTH 

eytes  and  necrosed  tissues.  All  analj'ses  of  pus  that  are  recorded  in 
the  literature  are  in  harmony  Avith  the  above  statements.  In  general 
the  analyses  consider  pus  as  composed  of  two  chief  portions,  the  pus- 
corpuscles  and  the  pus  serum.  As  is  to  be  expected,  the  composition 
of  pus-corpuscles  is  simply  that  of  a  large  mass  of  leucocytes,  which 
contain  minute  quantities  of  substances  taken  up  from  the  pus  serum 
by  absorption  and  phagocytosis.  The  old  analyses  of  pus-corpuscles 
by  Hoppe-Seyler  ®^  are  given  in  the  following  table : 

Table  I. 
Quantitative  Composition  of  Pus-cells   (in  1000  parts  of  the  dried  substance). 

I  n 

Proteins 137.62    1 

Nuclein 342.57    >  685.8.5  673.69 

Insoluble  bodies 205.66    J 

Lecithin ~l  i^qqq  75.64 

Fat         J"  '^^•^■^'^  75.00 

Cholesterol 74.00  .      .  72.83 

Cerebrin 51.99  )  iaqr-i 

Extractive  bodies 44.33  J  '      '  i"-o-* 

Mineral  Substances  in  1000  Parts  of  the  Dried  y'^ubstance. 

NaCI 4.35 

Ca,(P04)o 2.05 

Mg3(PO,)2 1.13 

FePO^ 1.06 

PO4 9.16 

Na 0.68 

K trace 

As  abnormal  constituents  of  the  leucocytes  contained  in  abscesses 
may  be  mentioned  glycogen,  fat  (from  phagocytosis  and  from  ''fatty 
degeneration"  of  the  leucocytes),  and  ''peptone"  (Hofmeister).^° 

Pus  serum  differs  from  blood-serum  chiefly  in  the  substances  added 
to  it  through  the  proteolytic  changes  that  occur  in  the  pus,  and  also 
in  that  it  has  lost  its  antiproteolytic  property,  containing  instead  free 
leueoprotease.  The  fibrinogen  that  escapes  from  the  vessels  into  sup- 
purating areas  becomes  so  altered  that  pus  will  not  coagulate,  even 
upon  addition  of  fibrin  ferment  (defibrinated  blood).  The  reaction 
of  the  serum  is  usually  slightly  alkaline,  becoming  strongly  alkaline 
if  much  ammonia  is  produced,  which  occurs  especially  if  there  is  sec- 
ondary infection  with  the  organisms  of  putrefaction.  Sometimes, 
liowever,  lipase  derived  either  from  bacteria  or  from  the  cells  causes 
the  splitting  of  sufficient  amounts  of  fatty  acids  from  the  fats  to  make 
the  reaction  acid ;  lactic  and  other  fatty  acids  are  also  sometimes 
formed.  Presumably  the  nature  of  the  infecting  organism  will  mod- 
ify the  reaction,  for  some  (e.  g.,  sfnphjflococcus)  cause  an  acid  forma- 
tion in  media,  while  otliers  (e.  g.,  pj/ocj/ancus)  cause  an  alkaline  reac- 
tion.    Hoppe-Seyler 's  analysis  of  pus  serinn  gave  the  following  re- 

68  Mod.-r'hom.  ITntersnoluinficn. 
TOZeit.  physiol    Cheni.,   1880    (4),  268. 


COMPOSITION  OF  PUS  279 

suits,  wliicli  show  no  considerable  deviation  from  the  composition  of 
blood  plasma,  except  in  an  increased  proportion  of  fatty  matter  and 
extractive  substances. 

Tarle  II 

Quantitative  composition  Plasma 

of  pus  serum  [normal ) . 

I  II  III 

Water 913.7  905.65  90S.4 

Solids 86.3  94.35  91.6 

Proteins 63.23  77.21  77.6 

Lecithin 1.50  0.56  "1 

Fat 0.26  0.29  }                1.2 

Cholesterol         0.53  0.87  J 

Alcohol  extractives      .      .      .  1.52  0.73  )                  .^ 

Water  extractives        .      .      .  11.53  6.92  (                 ^^ 

Inorganic  salts       ....  7.73  7.77  8.1 

Quantitatively  the  chief  abnormal  constituent  of  pus  serum  is  the 
so-called  " pyin"  of  the  older  writers,  which  is  nucleoprotein  de- 
rived from  the  decomposing  leucocytes,  and  hence  increasing  in 
amount  progressively  with  the  age  of  the  pus;  '^  it  is  characterized  by 
its  insolubility  in  acetic  acid.  The  same  substance  is  found  more 
abundantly  in  the  entire  pus,  on  account  of  the  presence  of  the  cells, 
and  when  treated  with  10  per  cent.  NaCl  solution  it  forms  a  stringy 
mass  which  was  formerly  called  "Rovida's  hyalin  substance."  Glu- 
cothionic  acid,  derived  from  the  leucocytes,  is  also  present  in  pus.'^- 
In  the  pus  serum  are  found  all  the  other  constituents  of  the  leuco- 
cytes, including  particularly  lecithin,  cholesterol,  fats  (and  soaps), 
cerebrin,  "jecorin, "  and  glycogen;  and  also  the  usual  components  of 
the  blood-serum  as  well  as  some  small  quantities  of  pigment  derived 
from  decomposed  red  corpuscles. 

The  products  of  autolysis  are  of  particular  interest,  and  they  are 
found  in  varying  amount,  but  usually  less  abundantly  than  might  be 
expected,  probablj'-  because  of  their  solubility  and  consequent  rapid 
absorption.  Albumoses  and  peptones  seem  to  be  constantly  present 
(Shattock).'^  The  common  occurrence  of  albumosuria  during  sup- 
puration presumably  depends  on  the  absorption  of  digestion  products 
from  the  pus,'^*  but  true  peptone  has  not  been  satisfactorily  identified 
in  the  urine.  Leucine  and  tyrosine  have  also  frequently  been  found 
in  pus,^^  but  Taylor  ''^  could  find  no  workable  traces  of  either  monoam- 

71  Strada.  Biochem.  Zeit..  1909    (16),  193. 

"2  Mandel  and  Levene,  Biochem.  Zeit..   1907    (4),  78. 

-3  Trans.  London  Path.  Soc,  1892   (43),  225. 

"4  Literature  on  albumosuria,  see  Yarrow.  Amer.  'Mod..  1903  (5).  452:  Elmer, 
ihirl..  1906  (11),  169;  Senator,  International  Clinics,  1905  (IV),  series  14.  p.  85. 
See  also  "Albumosuria,"  Chap.  xix. 

"5  Mfiller  (Cent.  inn.  Mod.,  1907  (28),  297)  recommends  the  tyrosine  reaction 
with  Millon's  reagent  as  a  means  of  difTerentiatiTi<j  tuberculous  from  ordinary 
pus.  the  former  not  pivinp  tlie  reaction  because  of  lack  of  leucocytic  enzymes; 
but  there  is  disagreoment  as  to  the  constancy  of  this  reaction  in  pus  (Dold,  Deut. 
med.  Woch.,  1908    (.34),  869). 

76  Univ.  of  California  Publications    (Pathol.),  1904    (1),  46. 


28P  IlsFLAMMATION,  REGENERATION,  GROWTH 

ino-  or  polyamino-acids  in  a  liter  of  pus,  which  may  depend  on  their 
havinof  been  eitlier  absorbed  or  transformed  into  ammoninm  com- 
pounds. From  the  nucleoproteins  purine  bodies  are  formed  and  may 
be  found  in  the  pus.  The  relation  of  the  purine  bases  to  local  leuco- 
cytosis  is  shown  by  Heile,"^  who  found  in  cold  tuberculous  abscesses 
a  proportion  of  purine  bases  equal  to  0.5  per  cent.,  in  similar  ab- 
scesses after  injection  of  iodoform,  1.57,  and  in  acute  suppuration, 
10.7.  Spermin  crystals  are  also  occasionally  found  in  old  pus  col- 
lections.'^^ Free  fatty  acids  and  volatile  fatty  acids,  such  as  butyric, 
lactic,"^  valerianic,  and  formic,  have  been  found.  Products  of  bac- 
terial activity,  such  as  bacterial  proteins  and  pigments  (e.  g.,  pyo- 
cyanin),  may  also  be  present.  It  is  probable  that  in  many  instances 
these  autolytic  products  are  bactericidal,  and  thus  help  to  terminate 
the  infection.  Direct  tests  have  shown  that  the  autolysate  of  fibrin  is 
bactericidal  for  staphylococci  and  streptococci.'^  See  also  discussion 
of  "Autolysis  of  Exudates"  (Chap.  iii). 

All  the  numerous  enzymes  of  the  blood  plasma,  the  leucocytes  and 
the  tissue-cells  are  present  in  pus.  Tluis  Achalme  *°  found  evidence 
of  the  presence  of  the  following  enzymes  in  pus :  proteolytic  en- 
zymes,®^ lipase  (splitting  monobutyrin),  diastase,  rennin  (coagulating 
milk),  gelatinase,  catalase,  and  oxidase,  the  last  being  very  abundant. 
These  seem  to  exist  chiefly  in  the  leucocytes,  the  pus  serum  being 
quite  free  from  them.  No  evidence  could  be  found  of  enzymes  act- 
ing on  amygdalin,  saccharose,  inulin,  or  lactose.  Fibrin  ferment  is 
said  to  be  absent  from  pus,  which  is  quite  surprising  in  view  of  the 
fact  that  this  enzyme  is  generally  considered  as  being  derived  chiefly 
from  the  leucocytes.  Presumably  the  bacteriolytic  "endolysins"  of 
the  leucocytes  are  also  present  in  pus. 

SPUTUM  S2 

The  chemistry  of  sputum  may  be  properly  considered  in  this  con- 
nection. In  reaction,  sputum  is  ordinarily  alkaline,  but  in  case  of 
marked  bacterial  decomposition  in  cavities  the  reaction  may  become 
acid.  Its  specific  gravity  varies  from  1.008  to  1.026,  usually  varying 
directly  witli  the  number  of  leucocytes ;  the  average  specific  gravity 
is  about  1.013.  The  greenish  color  frequently  observed  depends  gen- 
erally upon  blood-pigment   (except  in  case  of  icterus),  although  in 

77  See  Williams,  Boston  IMcd.  and  Sip-ct.  Jour.,  1901    (14,5),  355. 

78  d-lactic  acid  is  a  constant  constituent  of  pus  from  the  pleura  (Ito,  Jour. 
Biol.  Chem.,  11116    (26),  173). 

70  Bilancioni,  Arch,  di  Farmac<d.,  1911    (11),  491. 
soCompt.  Rend.  Soc.  Biol.,  1S99    (.51),  56S. 

81  Concerning  proteolytic  enzvmes  of  pus  see  Opie,  Jour.  Expor.  Med..  1906  (8), 
410. 

82  Complete  hihlio^aphv  given  hv  Ott,  "Cliem.  Patiiol.  dcr  Tubprc,"  Berlin, 
1903;  lalk,  Krgehnisse  Plivsiol.,  1910  (9),  406:  Plesch,  Handb.  d.  Biochem.,  1908 
(HI  (1)   ),  7. 


SPUTUM  281 

some  instances  the  pigment  is  of  bacterial  origin.  Renk  *^  has  stud- 
ied the  proteins  of  sputum  with  special  reference  to  the  loss  of  pro- 
tein to  the  body  and  its  relation  to  cachexia.  In  three  patients  (con- 
sumptives) studied,  the  daily  amount  of  sputum  of  two  averaged  145 
grams  for  each;  for  the  third  it  was  82  grams.  This  contained 
(average)  5  to  6  per  cent,  of  solids;  including  mucin,  2-3  per  cent.; 
protein,  0.1-0.5  per  cent. ;  fat,  0.3-0.5  per  cent. ;  ash,  0.8-0.9  per 
cent.  The  daily  loss  of  nitrogen  was  0.75  gram,  which  equals  about 
6  per  cent,  of  the  total  daily  nitrogen  output  of  persons  under  condi- 
tion of  starvation.^*  Wanner  *^  found  characteristic  variations  in  the 
amount  of  protein  in  sputum  from  different  conditions,  as  follows : 
in  bronchitis  the  amount  of  protein  is  very  small ;  in  bronchiectasis 
protein  is  present,  but  the  amount  of  uncoagulable  nitrogen  (due  to 
autolysis)  is  relatively  large;  in  phthisis  as  well  as  in  bronchiectasis 
the  amount  of  protein  does  not  exceed  1  per  cent.,  in  pneumonia  it 
may  reach  3  per  cent.,  but  it  is  highest  in  pulmonary-  gangrene.  Any 
protein  content  that  causes  more  than  a  slight  turbidity  on  boiling  in- 
dicates an  inflammation ;  e.  g.,  in  case  of  doubt  between  a  diagnosis  of 
I>neumonia  and  infarct  a  high  protein  content  speaks  for  the  former. 
Rogers  ^^  stated  that  the  sputum  in  every  case  of  tuberculosis  shows 
albumin,^*'-''  but  this  has  been  questioned,  especially  as  to  chronic  or 
quiescent  cases. ^^  Albumin,  or  better,  coagulable  protein  is  also  pres- 
ent in  the  sputum  of  patients  with  pulmonary  edema  and  pleurisy. 
According  to  Works  ^^  in  active  tuberculosis  there  is  usually  0.2  per 
cent,  or  more  of  coagulable  protein  in  the  sputum.  The  mucin  of 
sputum  yields  33.6  per  cent,  of  glucosamin  when  split  with  HCl,  which 
gives  an  index  of  the  quantity  of  mucin ;  this  is  highest  in  chronic 
bronchitis  and  lowest  in  pneumonia  and  phthisis.  Kossel  found  0.1- 
0.33  gm.  of  nucleins  in  the  sputum  daily. 

The  following  table  by  Bokay  (taken  from  Ott)  gives  the  pro- 
portion of  the  organic  constituents  of  sputum  in  parts  per  thou- 
sand : 

On  account  of  the  digestion  of  the  exudates  by  the  leucocytes,  spu- 
tum contains  proteoses,  peptones,  and  amino-acids,  generally  in  pro- 
portion to  the  richness  of  the  exudate  in  leucoej-tes;  they  are,  there- 

ssZeit.  f.  Biol.,  187.5   (11).  102. 

84Plesch  (Zeit.  exp.  Path.  u.  Ther..  1906,  Bd.  iii.  .Tuly)  found  that  4.8  per 
cent,  of  all  the  absorbed  calories  were  lost  in  the  sputum  in  an  advanced  case  of 
phthisis.  Under  similar  conditions  the  amount  of  salts  excreted  by  tlie  sputum 
mav  equal  or  exceed  that  in  the  urine  ( Falk,  loc.  cit. ) . 

ssDeut.  Arch.  klin.  Med..  1003    (75).  .347. 

ssPresse  Mfd.,  1910  (18),  289;  1911  (19).  409;  also  Ganz  and  Hertz,  ibid., 
1911  (19),  41;  Kaufmann,  Beitr.  Klin.  d.  Tuberk,  1913  (26),  269;  Hempel- 
Jfirgensen.  ibid.,  p.  392. 

86a  RevieAv  by  Cocke.  Amer.  Jour.  Med.  Sci..  1914   (148).  724. 

8"  Fischberfr  and  Felberbaum,  Medical  Record,  Oct.  21,  1911;  Acs-Xagv,  A^'ien. 
klin.  Woch.,   1912    (25),  1904. 

88  Jour.  Amer.  Med.  Assoc,  1912   (59),  1537. 


282 


IXFLAMMATIOX,  liEGEXEh'ATIOX,  GROWTH 


Table  III. 

Bronchitis 

Phthisis, 

Phthisis, 

in 

Fibroid             early  in 

Phthisis, 

Phthisis, 

ad- 

typhoid 

phthisis               apex 

cavities 

advanced 

vanced 

Fatty  acids  as  fat 

0.224 

0.845             0.462 

2.468 

3.468 

9.725 

Free  fatty  acids  . 

trace 

0.184             0.521 

0.370 

0.307 

0.902 

Soaps    .... 

traces 

0.380            0.430 

0.537 

0.516 

3.973 

Cholesterol 

traces 

0.4                 1.617 

0.172 

1.160 

0.141 

Lecithin 

traces 

traces           1.543 

1.165 

1.245 

Xuclein 

traces 

0.102 

0.260 

0.489 

Protein 

0.898 

2.040 

4.430 

3.455 

5.115 

fore,  most  abundant  in  pneumonia.  Simon  '^^  found  considerable  al- 
bumose  in  phthisical  sputum,  but  no  nucleohiston  or  free  histon. 
In  febrile  cases  of  tuberculosis  the  amount  of  albumose  may  exceed 
the  coagulable  albumen,  which  rarely  exceeds  one  per  cent,  of  the 
moist  weight.""  Staffregen,  however,  could  find  no  true  peptone  in 
phthisical  sputum,  but  Stadelmann  -'^  found  that  such  sputum  con- 
tained enzymes  hydrolyzing  fibrin,  and  attributed  this  largely  to 
bacteria.  Probably  most  of  the  enzjones  present  in  sputum  come 
from  the  leucocytes.  In  the  early  stage  of  pneumonia  the  sputum 
has  no  proteolytic  action,  presumably  because  inhibited  by  the  large 
amount  of  serum  present ;  but  with  resolution  active  proteolji:ic  prop- 
erties appear  (Rittorf).^-  In  tuberculosis  sputum  the  tryptic  and 
antitryptic  properties  fluctuate,  and  lipase  is  absent  (Eiselt).®^ 
Pneumonic  sputum  before  the  crisis  has  but  slight  action  on  peptids, 
but  acquires  marked  peptolytic  activity  thereafter.^*  Most  sputa 
contain  enzymes  splitting  casein  and  polj'peptids.''*''  Sputum  may 
contain  indole,  derived  either  from  the  putrefying  proteins  or  ex- 
creted from  the  blood."*'' 

The  amount  of  fats  seems  to  depend  directly  upon  the  number  of 
pus-corpuscles  and  the  age  of  the  pus  (i.  e.,  the  amount  of  fatty  de- 
generation). Jacobson  found  from  0.08  to  1.6  grams  of  fatty  matter 
per  day,  containing  on  an  average  14.76  per  cent,  of  soaps,  15.79  per 
cent,  of  higher  fatty  acids,  0-10  per  cent,  of  water-soluble  fatty  acids, 
13.58  per  cent,  lecithin,  and  10.49  per  cent,  cholesterol. 

As  to  the  inorganic  substances.  Bamberger  found  two  types  of  spu- 
tum, catarrlial  and  inflammatory.  In  tlie  inflammatory  there  is  a 
deficiency  in  alkali  phosphate,  SO;,- constitutes  more  than  8  per  cent. 

Na  O  .        15 


of  the  salts,  and  the  ratio, 


KO 


equals 


41 


the  alkali  phosphates  constitute  10-14  per  cent.,     -rr^ 


In  catarrhal  sputum 

Na  O  31 

^,  and  the 


KO 


89  Arch.  exp.  Path.  u.  Pharm.,  1903    (49),  449. 
ooProrok.  Miinoh.  mod.  Wodi..  1909    (56).  2053. 
fi  Zeit.  klin.  Med..  1SS9   (16).  128. 
02Deut.  Arch.  klin.  Mod.,   1907    (91).  212. 
93Zoit.  klin.  Mod.,  1912    (75),  91. 

04  Ahdorlialdon.  Zoit.  phvsiol.  C'liom..   1912    (78),  344. 
n4a:Maliwa,  Dout.  Aroh.  klin.  Med.,  1914   (115),  407. 
94b  Binda  and  Cassarini.  Oaz.  Med.  Ital.,  1913    (64), 


461. 


J'h'<)IJI/:U.\T/(>\  AM)  h'KdENERATION 


283 


SO;,  is  from  0.6-1.2  per  cent.  Chlorine  is  about  the  same  in  botli 
forms.  These  differences  are,  however,  not  as  constant  as  Bamberger 
believes,  aecordino-  to  several  later  inv('sti<i'ati()ns.  The  i-cstilts  of  his 
analyses  are  shown  in  the  following  table : 

Table  IV 


Water 

Orj^anic  matter 

Inorganic   salts 

One  hundred  parts  of  the  salts  contain: 

Chlorine 

SO, 

P260 

K2O 

NaoO         

Calcium  phosphate 

Iron  pliosphate 

jNIagnesiuni  phosphate 

Ca  and  Mg  carbonate  and  sulpliate 

Silicic  acid     .  


Chronic 
phthisis 


!)4.5.-> 
4.(i7 
0.78 


35.78 

0.70 

13.05 

24.07 

27.1)0 

1.G3 

0.09 

1.20 

1.74 

0.0 


Acute 
phthisis 


93.38 
6.88 
0.74 


33.40 

0.80 

14.15 

19.99 

31.09 

4.32'- 

0.14 

0.22 

0.3 


PROLIFERATION  AND  REGENERATION 

The  factors  that  incite  cells  to  proliferate,  as  well  as  those  that 
cause  the  cessation  of  proliferation  after  it  has  once  started,  are  too 
entirely  unknown  to  permit  of  speculation  as  to  their  exact  nature. 
It  seems  probable,  however,  that  they  are,  as  Ziegler  says,  "identical 
with  the  stimuli  which  excite  or  increase  functional  and  nutritive 
activity,"  and  these  are  certainly  in  many  instances  of  chemical  na- 
ture. Thus  the  application  of  various  irritating  substances  in  not 
too  concentrated  a  form  (e.  g.,  painting  the  skin  with  iodin)  may 
lead  to  proliferation  without  causing  discernible  degeneration  of  the 
cells.  Mallory's^"'  observations  on  the  phenomena  of  proliferation 
and  phHgocvlT)susjjhowJii^rit  the  same  baj-terial  products  whicli  destroy 
the  cells  when  concentrated,  when  sufficiently  dilute  cause  prolifera- 
tion  of  similar  cells.  Carnot  and  Lalievre ""  have  obtained  evidence 
that  actively  growing  kidney  tissue,  whether  fetal  or  adult  regener- 
ating kidney,  contains  something  which  is  capable  of  stimulating 
growth  of  renal  epithelium  when  injected  into  other  animals.  (The 
importance  of  this  observation  calls  for  its  corroboration,  but  no  repe- 
tition of  the  work  is  known  to  us.)     Many  other  instances  of  prolifera- 

^•5  Including  magnesium. 

98  Jour.  Exp.  Med.,  1900   (5),  15. 

97  Arch.  M6d.  Exper.,  1907   (19),  388. 


284  JXFLAMMATIOX,  REGENERATION,  GROWTH 

tiou  in  response  to  chemical  stimuli  might  be  cited,  but  in  nearly  all 
cases  it  is  extremely  difficult  to  determine  that  the  proliferation  is 
not,  after  all,  reparative  in  compensation  for  degenerative  changes, 
and,  therefore,  possibly  obeying  some  other  biological  law  than  that 
of  a  simple  reaction  to  a  chemical  stimulus. 

Although  proper  nutrition  is  necessary  for  cell  proliferation,  yet  it 
does  not  seem  that  excessive  nourishment  can  lead  to  excessive  cell  mul- 
tiplication, or  by  itself  cause  cell  proliferation  to  take  place.  Oxygen 
and  certain  inorganic  salts  are  essential  for  cell  division  even  in  the 
lowest  forms,  and  among  such  simple  organisms  as  sea-urchins  and 
certain  other  marine  forms  segmentation  of  the  unfertilized  ova  may 
be  incited  by  changes  in  osmotic  concentration,  leading  eventualh*  to 
formation  of  perfect  larvje  (J.  Loeb,  et.  al.).^^  In  lower  animals 
verj'  dilute  solutions  of  alkalies  stimulate  the  rate  of  cell  growth,  and 
somewhat  higher  concentrations  cause  extremely  irregular  cell  division ; 
in  mammals  the  feeding  of  alkalies  causes  great  wasting  as  if  through 
cell  stimulation.^  The  products  of  nuclein  hydrolysis  are  said  to 
stimulate  cell  growth.-  Potassium  salts  seem  to  be  particularly  im- 
portant for  proliferating  cells,  and  Beebe  and  also  Clowes  and  Fris- 
bie  ^  have  found  that  actively  growing  malignant  tumors  are  rich 
in  potassium  and  poor  in  calcium,  whereas  in  slow-growing  tumors 
the  reverse  is  the  case.  Dennstedt  and  Rumpf  *  also  found  that  in 
hypertrophy  of  the  heart  the  amount  of  potassium  is  increased,  while 
in  chronic  degeneration  of  the  myocardium  the  calcium  and  mag- 
nesium are  usually  increased.  The  proportion  of  nitrogen  in  the  dif- 
ferent parts  of  the  heart  is  not  changed  during  hypertrophy  (Benee),^ 
but  the  amount  of  NaCl  is  much  increased  in  hypertrophy." 

Chemical  studies  of  proliferation  are  lacking,  except  in  regard  to 
the  development  of  the  embryo,  etc."'"^  New  tissues  difiPer  from  adult 
tissues  in  having  a  large  proportion  of  water,  and  in  having  a  larger 
proportion  of  the  "primary"  cell  constituents  and  a  smaller  propor- 
tion of  the  various  secondary  constituents,  since  these  last  are  largely 
products  of  the  activity  of  the  adult  cell.  Of  the  primaiy  constitu- 
ents, the  proportion  of  the  nucleoproteins  is  particularly  high,  and  a 
number  of  interesting  facts  concerning  the  nucleoproteins  in  cell  di- 
vision have  been  determined.  IMost  important,  perhaps,  are  the  clas- 
sical observation  of  INIiescher,  who  found  that  during  the  migration 
of  salmon  up  stream  to  the  spawning  grounds,  during  which  time  no 
food  is  taken,  the  proteins  of  the   muscular  tissue  become  largely 

08  Sep  J.  Loob,  Studios  in  Conoral  Plivsiolofrv,  Chioapo.  1005. 

1  Moore  et  al.,  Rioolicm.  Jour..  1900    h),  2\H:   1012    (6),  162. 

2  Calkins  et  al..  Jour.  Infect.  Di.s.,  1912    (10),  421. 
8  See  "Tumors,"  Cliap.  xvii. 

4Zeit.  klin.  Med.,  190.5    (58).  84. 
BZeit.  klin.  Med..  1908    (06).  441. 
«  Rzentkowski,  ihid.,  1910    (70),  3.37. 

"a.  l.iterature  on  the  ehemistry  of  firowtli  piven  liy  Aron,  llandliueli  d.  Bioeliem., 
Ergiinzungsband,  1913. 


CHEMICAL  /?.hS7<S'  OF  (IROWTll  A\D  REl'Mli.     'TIT  [  M I  \JJ,S"     285 

transformed  into  the  protamin  t3^pe  of  protein  (characterized  by  con- 
taining hirji^e  proportions  of  tlir  ixilyamiiio-acids,  such  as  arj^inine, 
histidine,  and  lysine)/  which  unite  with  nucleic  acids  to  form  the 
abundant  nucleoprotein  of  the  spermatozoa  and  ova.  Whether  such  a 
transformation  of  proteins  occurs  in  mannmdian  cells  during  cell 
multiplication  cannot  be  stated,  but  certainly  from  some  source  an 
additional  supply  of  nucleoprotein  is  derived.  Developing  sea  urchin 
eggs  synthesize  great  quantities  of  nucleoprotein,^''  even  when  in  a 
solution  free  from  phosphates,  and  here  the  only  available  source  for 
the  phosphoric  acid  of  the  nucleins  would  seem  to  be  the  lecithin  of 
the  egg  (J.  Loeb).  The  nucleoproteins  during  karyokinesis  undergo 
a  chemical  change  in  that  they  become  of  a  more  acid  type  (presum- 
ably through  splitting  off  of  part  of  the  proteins  from  the  nucleic 
acid),  which  results  in  the  characteristic  increase  in  affinity  for  basic 
dyes,  and  the  increased  negative  charge  which  is  easily  demonstrated.® 
This  suggests  the  participation  of  an  enzyme  in  the  process  of  karyo- 
kinesis, just  as  there  seems  to  be  in  the  production  of  pycnosis  in  de- 
generating cells,  but  there  seems  to  be  no  conclusive  evidence  on  this 
point.  Gies "  could  find  no  enzyme  in  spermatozoa  that  incites  cell 
division  in  the  ova  of  sea-urchins  (Arhacia).  The  fertilization  of 
eggs  makes  them  more  permeable  to  ions,^"^  which  possibly  determines 
many  of  the  subsequent  changes. 

In  metaplasia  we  have  what  may  be  interpreted  as  a  chemical  alter- 
ation due  to  mechanical  stimuli,  e.  g.,  the  fonnation  of  keratin  by  cells 
that  ordinarily  do  not  do  so ;  the  deposition  of  calcium  salts  and  oste- 
oid transformation  of  connective  tissues  in  rider's  bone,  etc.  That 
such  is  the  case,  however,  cannot  be  positively  stated  from  the  evidence 
at  hand. 

CHEMICAL  BASIS  OF   GROWTH  AND   REPAIR.     '"VITAMINES"  i- 

We  do  not  know  just  what  substances  are  necessary  to  maintain 
individual  cells  in  normal  conditions,  what  are  needed  to  stimulate 
them  to  multiplication,  or  what  elements  they  require  to  permit  them 
to  multiply,  but  it  has  been  learned  that  certain  definite  materials  are 
required  by  the  organism  as  a  whole.  It  is  not  sufficient  that  a  given 
number  of  calories,  with  a  certain  quantity  of  proteins,  carbohy- 
drates, fats  and  salts  be  supplied ;  it  is  essential  that  certain  specific 
constituents  be  provided  among  these  foodstuffs.     The  proteins  must 

7  Concerning  protamins,  see  resume  bv  Kossel,  Biocliem.  Ceiitr..  1000  C5),  1 
and  33. 

Ta  Xot  accepted  bv  I^Tasing,  Zcit.  plivsidl.  Clieiii..  1010   (fi7),  IHl. 

'^See  Gallardo,  Arch.  Entwickl.  Organ.,  1000  (2S),  12.i:  Pentiinalli,  U>i<1..  1012 
(34).  444. 

oAmer.  .Tour.  Phvsiol..  1001    ffi).  .54. 

11  See  IMcClendon',  Carnegie  Inst.  Publ..  1014,  Xo.   1S3. 

12  See  Mendel,  "Nutrition  and  Growth,"  Harvev  Society  Lectures.  1014- I.t: 
Amer.  Jour.  Med.  Sci.,  1917   (153),  1. 


286  JXFLAMMATJOX.  RECEy ERATIOy .  aROWTH 

not  only  provide  a  sufficient  amount  of  nitrogen,  but  they  must  also 
provide  certain  specific  amino-acids,  as  has  been  especially  demon- 
strated by  the  investigations  of  Osborne  and  JMendel.'^  Apparentlj' 
the  presence  of  some  of  the  simj)le  straight-chain  amino-acids  can  be 
dispensed  with  (c.  g.,  g-lycine),  and  the  animal  will  grow  and  thrive 
if  other  nutritive  supplies  are  adequate,  but  certain,  at  least,  of  the 
more  complex  cyclic  amino-acids  must  be  provided.  Furthermore, 
the  requirements  for  growth  (quantitatively  speaking  at  least),  seem 
to  be  sometliing  more  than  the  requirements  for  mere  preservation  of 
health  and  equilibrium,  for  it  was  found  that  animals  could  live  and 
preserve  nitrogen  equilibrium  when  the  protein  of  the  diet  furnished 
at  most  small  quantities  of  lysine,  but  young  animals  were  unable  to 
grow  with  such  a  restricted  supply  of  this  amino-acid.  If  lysine  was 
added  to  the  defective  protein  (gliadin  from  wheat)  the  animal  would 
then  be  able  to  grow  at  a  normal  rate.  Of  particular  importance  is 
the  fact  that  animals  can  be  kept  in  a  stunted  condition  on  such  a 
deficient  diet  until  they  have  reached  an  age  at  which  normally  all 
growth  would  have  long  since  ceased,  and  then  when  supplied  with 
sufficient  lysine  they  will  begin  to  grow  and  continue  until  full  size  is 
reached.^*  This  last  observation  proves  that  growth  is  not  condi- 
tioned by  age,  and  that  we  do  not  stop  growing  because  a  certain  age 
is  reached :  the  capacity  for  growth  may  remain  latent  and  capable 
of  exhibiting  itself  when  proper  conditions  are  furnished.  But  no 
amount  of  any  amino-acid  will  cause  a  fully  grown  animal  to  grow 
any  more,  so  it  would  seem  that  the  capacity  for  growth  becomes  ex- 
tinguished when  it  has  been  utilized  to  a  certain  fixed  extent,  and  re- 
mains potent  until  it  has  been  completely  utilized. 

If  the  only  protein  furnished  contains  no  tryptophane  the  animal 
cannot  maintain  itself  and  slowly  loses  weight  until  it  dies,  unless 
tryptophane  is  supplied.  If  zein  from  corn,  which  yields  neither  h'- 
sine  nor  tryptophane,  is  the  sole  protein,  then  the  animal  cannot  grow 
unless  both  lysine  and  tryptophane  are  added  to  the  diet.  That  the 
pure  isolated  amino-acids  can  meet  the  deficiencies  when  added  to  the 
imperfect  protein  ration,  demonstrates  that  proteins  ser^'e  for  food  as 
amino-acids,  and  not  as  larger  complexes. 

Not  only  must  the  proteins  present  certain  essential  chemical  com- 
pounds to  the  living  and  growing  organism,  but  also  an  adequate 
su])])ly  of  the  essential  inorganic  salts  and  certain  other  less  well  de- 
fined substances  are  also  necessary  to  permit  of  maintenance,  growth 
and  repair.  It  has  long  been  recognized  clinically  that  certain  dis- 
eases, notably  scurvy,  may  result  from  tlu>  absence  of  some  essential 
food  supply.  More  recently  other  diseases  have  been  proved  or  sus- 
pected of  having  a  similar  cause,  and  the  study  of  one  of  these,  beri- 

13  Series  of  papers  in  Jour.  Biol.  Clioiii..   1!)12,  ct  scf/. 
^•».Tnnr.  P.inl.  Diem.,   1915    (23),  4:W. 


\nA.\u\Ei^  287 

beri,  lias  led  to  a  closer  approximation  to  the  nature  of  the  food  essen- 
tials concerned.  This  disease  seems  to  result  from  the  use  of  pol- 
ished rice  as  the  chief  constituent  of  the  diet,  and  can  be  checked  by 
feeding  unpolished  rice,  or  rice  polishings,  or  even  extracts  of  rice 
polishiugs.  A  somewhat  similar  condition  may  be  produced  readily 
in  birds  by  feeding-  polished  rice,  the  chief  feature  being  a  severe 
neuritis,  which  is  relieved  with  remarkable  rapidity  by  supplying  the 
food  deficiency.  This  experimental  neuritis  of  fowls  {polyneuritis 
gallinarum)  has  served  as  a  valuable  means  of  study  of  diseases  of 
this  class,  and  led  to  the  demonstration  that  not  only  extracts  of  rice 
polishings,  but  also  many  other  food  materials,  contain  the  essential 
materials  without  which  health  cannot  be  maintained.  One  of  the 
earl}'  investigators  of  this  subject,  Casimir  Funk,^"'  gave  to  "the 
hitherto  unrecognized  essential  dietary  factors"  the  name  "vitamines, " 
which,  in  spite  of  certain  logical  objections,  has  been  generally  adopted. 
Although  so  essential  for  life  the  amount  required  is  very  small,  for 
whole  rice  is  said  to  contain  not  over  0.1  gm.  per  kilo.,  and  perhaps 
much  less,  of  the  active  substance.  Besides  beriberi,  the  following 
may  be  considered  as  of  the  nature  of  ' '  deficiency  diseases ' ' :  Scurvy 
and  infantile  scurvy  (Barlow's  disease)  and  possibly  pellagra,  rick- 
ets and  tetany.  Furthermore,  the  "vitamines"  are  necessary  for 
growth,  no  matter  how  much  and  what  proteins,  carbohydrates  and 
fats  are  supplied. 

McCollum  '■^  has  summarized  the  evidence  that  two  classes  of  sub- 
stances are  necessary  for  maintenance,  the  more  important  in  pre- 
venting neuritis'  being  water-soluble,  although  for  growth  to  occur  an 
unidentified  lipoid-soluble  substance  is  essential.  As  yet  the  exact 
identity  of  the  active  agents  in  water  or  lipoid  solutions  has  not  been 
determined,  but  the  pursuit  is  being  gradually  brought  closer  to  the 
goal.  Funk  believed  them  to  be  pyrimidine  derivatives.  AVilliams 
and  Seidell  ^^  have  found  that  hydroxypurines  have  marked  anti-neu- 
ritic  effects,  and  they  suggested  that  an  isomer  of  adenine  is  responsible 
for  the  antineuritic  action  of  yeast  extracts.  Later  Williams  ^''"^ 
found  an  active  hydroxypyridene,  and  suggested  that  the  curative 
properties  of  yeast  and  rice  polishings  may  be  due  to  an  isomeric  form 
of  nicotinic  acid.  The  lipoid-soluble  "vitamines"  seem  to  be  especially 
abundant  in  butter,  egg  yolk,  and  cod  liver  oil,  which  presumably  ac- 
counts for  the  commonly  accepted  values  of  these  particular  fats. 
Why  the  vitamines  are  essential  and  how  they  act  is  unknown.  Ved- 
der  ^^  has  suggested  that  the  antineuritic  vitamine  is  essential  for 
growth  and  repair  of  the  nervous  tissue,  and  in  its  absence  normal 

15  See  Ergeb.  Phvsiol.,   1913    (13),  12.i,  for  review  of  liis  work. 
ifiJour.  Biol.  Chem.,   1916    (24),  491. 

17  Jour.  Biol.  Chem.,  1916    (26),  431. 
17a  Jour.  Biol.  Chein.,  1917    (29).  49"). 

18  Jour.  Amer.  Med.  Assoc,  1916    (67),  1494. 


288  INFLAMMATIOy,  REaENERATlOX,  GROWTH 

Avear  cannot  be  made  good.  It  is  probable  that  more  than  one  vita- 
mine  is  neeessaiy  for  maintaining  normal  conditions,  and  deficiency 
of  one  causes  beriberi,  of  another  scurvy,  for  some  dietaries  lead  to 
one  disease  and  some  to  the  other. 


CHAPTER    XI 

DISTURBANCES  OF  CIRCULATION  AND  DISEASES 
OF  THE  BLOOD 

THE  COMPOSITION  OF  THE  BLOOD 

The  function  of  the  blood  being-  to  maintain  an  equilibrium  in 
the  temperature,  chemical  composition  and  osmotic  pressure  between 
all  parts  of  the  body,  it  follows  that  it  is  never  of  exactly  the  same 
composition  in  any  two  places  or  at  any  two  times.  To  the  extent 
that  every  tissue  is  continually  giving  off  something  to  the  blood,  we 
may  consider  that  every  organ  is  a  factor  in  its  formation,  .and  as  a 
result  of  this  multiplex  origin  of  the  blood,  the  substances  it  may  con- 
tain are  beyond  enumeration.  There  are  probably  but  few  chemical 
substances  occurring  in.  the  tissue-cells  that  do  not  also  occur  iii 
greater  or  less  amount  in  the  blood.  In  addition  to  these  there  are 
also  the  substances  characteristic  of  the  blood  itself,  besides  a  host 
of  substances  of  unknown  nature,  apparently  manufactured  in  re- 
sponse to  the  stimulation  of  substances  entering  the  body  from  out- 
side ;  for  we  find  that  the  blood  of  every  adult  individual  contains 
substances  that  make  him  immune  to  a  multitude  of  diseases  that  he 
has  had  in  childhood,  as  well  as  substances  that  in  later  life  protect 
him  to  a  greater  or  less  degree  from  infection  by  such  organisms  as 
the  colon  bacilli  of  his  intestine,  the  pneumococci  and  streptococci 
in  his  throat,  etc.  We  have  learned  of  these  defensive  substances 
within  very  recent  times,  and  also  of  the  "  antienzymes "  that  possi- 
bly protect  the  blood  from  the  digestive  enzymes  of  the  body  cells. 
Wliat  other  substances  of  importance  we  may  yet  find  in  the  blood  is 
an  open  question.  There  are  no  apparent  limits  to  the  possibilities 
of  the  study  of  the  blood,  for  it  represents  a  little  of  every  organ, 
and  much  that  is  characteristic  of  itself. 

In  discussing  briefly  the  substances  that  have  been  isolated  from 
the  normal  blood,  before  considering  the  changes  that  occur  in  it 
during  pathological  conditions,  we  may  roughly  divide  the  blood  into 
the  formed  elements  and  the  plasma  in  which  they  are  suspended. 

THE  FORMED  ELEMENTS. — By  weijjht,  tlie  red  corpiisclos  constitute  from  40 
to  50  per  cent,  of  tlie  l)lo(id,  the  percentage  varying  under  difl'erent  conditions, 
while  the  total  weight  of  tlie  leucocytes  and  platelets  is  insignificant.  The  hemo- 
globin constitutes  from  Sii  to  04  per  cent,  liy  weight  of  tlie  solids  of  the  red  cor- 
puscles, but  the  physical  and  chemical  relations  that  it  bears  to  the  stroma  of  the 
corpuscles  are  as  yet  undetermined  (see  ''Hemolysis").  Of  tlie  remainin£r  constit- 
uents of  the  corpuscles,  from  .5  to  12  per  cent,  consist  of  proteins,  probably  chiefly 
globulins  and  nucleoproteins;  0.3  to  0.7  per  cent,  of  lecithin;  and  about  0.2  to  0.3 
19  289 


290  DISTURBANCES    OF    CIRCULATION 

per  cent,  of  cholesterol  (Hoppe-Seyler) .  The  outer  coat  of  tlie  red  corpuscles 
does  not  seem  to  be  equally  permeable  for  all  substances,  and  tlierefore  we  find 
the  composition  of  the  fluid  portion  of  the  cell  quite  different  from  tliat  of  the 
plasma  about  it.  The  salts  of  the  corpuscles  consist  largely  of  potassixun  ])hos- 
phate,  a  little  sodium  chloride,  some  magnesium,  but  no  calcium.i  which  is  quite 
different  from  their  proportion  in  tlie  plasma.  Probably  many  of  tlie  other  con- 
stituents of  the  plasma,  especially  urea,  penetrate  the  red  corpuscles  to  a  greater 
or  less  degree,  but  most  of  them,  particularly  the  sugar,  remain  chiefly  in  the 
plasma. 

Hemoglobin,  the  most  characteristic  constituent  of  all  the  heterogeneous  com- 
ponents of  the  blood,  is  a  compound  protein,  and  probably  exists  combined  with 
some  other  constituent  of  the  corpuscle,  most  probably  the  lecitliin.  It  splits 
up  readily  into  a  protein,  glohin,  and  an  iron-cont^iining  substance,  he^nochromo- 
gen,  which  readily  takes  up  oxygen  to  form  hematin.  Only  about  4  to  5  per  cent, 
of  the  hemoglobin  is  heniochromogen,  and  iron  constitutes  but  about  0.4  per  cent. 
Hematin  may  be  further  split  up  into  other  substances,  which  will  be  considered 
in  the  discussion  of  "Hemorrhage." 

The  leucocytes  consist  chiefly  of  nucleoproteins,  with  probaldy  some  globulin, 
and  they  also  contain  glycogen,  lecithin,  and  cholesterol.  The  hiood-pUitclrts  are 
believed  to  Ik^  largely  nucleoprotein,  but  little  is  known  of  tlicir  actual  composi- 
tion;   miiTocliciuical  examination  shows  no  evidence  of  either  fat  or  glycogen. 2 

BLOOD  PLASMA  differs  from  blood-serum  in  that  the  latter  is  formed  from  the 
former  through  the  conversion  of  the  fibrinogen  into  fibrin.  Serum,  therefore, 
contains  no  fibrinogen,  but  more  fibrin  ferment:  otherwise  it  is  practically  the 
same  as  the  plasma.  It  is  well  for  us  to  appreciate  that  the  blood  is  funda- 
mentally a  tissue,  with  its  more  solid  structural  elements  lying  in  a  protoplasm, 
the  plasma,  somewhat  more  dilute  than  the  protoplasm  of  other  tissues  but  in 
other  respects  much  the  same. 

Proteins. — Fibrinogen,  has  the  general  properties  of  a  globulin,  with  also  a 
peculiar  tendency  to  go  into  the  insoluble  form,  fibrin.  (Tliis  process  will  be 
discussed  imder  "Thrombosis.")  In  the  plasma  are  also  other  globulins.-'a  one 
soluble  in  water  (pseudo-glolmlin) ,  the  other  insoluble  in  water  (euglobulin) . 
Serum-albtimin,  another  protein  of  the  plasma,  probably  consists  of  two  or  more 
varieties  of  albumin.  There  are  also  nucleoproteins  (prothrombin)  and  non- 
coagulable  proteins,  which  being  poorly  understood  have  been  variously  considered 
as  glycoproteins,  or  mucoids,  or  albumoses.  The  serum  proteins  seem  to  be  closely 
related  to,  or  compovmded  with,  the  lipins  of  the  plasma. 

Other  Constituents. — The  fnt  of  the  plasma  varies  much  according  to  tlie  time 
which  has  elapsed  after  tlie  taking  of  f^od ;  in  fasting  animals  it  amounts  to  from 
0.1  to  0.7  per  cent.  The  sugar  fluctuates' less,  being  normally  about  0.1  pei-  cent., 
while  the  urea  has  been  estimated  at  0.05  per  cent.  ]\Tost  of  the  sugar  is  dex- 
trose; but  probably  there  is  some  levulose,  possibly  some  pentose  and  otlier  forms, 
and  possibly  also  sugar  combined  with  lecithin  (jecnrin)  or  other  substances. 
Soaps,  cholesterol,  and  lecithin  also  exist  free  in  the  plasma. 

Plasma  difTers  strikingly  from  the  corpuscles  in  that  its  inorganic  substances 
are  chiefly  sodium  and  chlorine,  while  potassium  and  phosphoric  acid  are  almost 
entirely  absent.  Another  imjiortant  fact  is  that  wIumi  the  plasma  is  combusted, 
the  acid  radicals  remaining  do  not  suffice  to  balance  the  bases,  indicating  that 
much  of  tlie  inorganic  bases  is  joined  with  organic  sulistances.  probably  as  ion- 
protein  compounds.  The  alkali  joined  to  tlie  protein  is  non-diffusible,  and  con- 
stitutes aliout  five-sixths  of  tlie  total  alkali. 

The  concentration  of  the  electrolytes  of  the  blood  has  been  determined  ^ly 
ascertainiiiir  iht>  lowering  of  the  f-ce/jim-iioint,  which  in  human  Idood  averages 
about  0..52(i° ;  this  corresponds  closcb-  to  the  efi'ect  of  a  salt  solution  of  0.0  per 
cent,  strength.  About  three-fcnirtlis  of  the  dissolved  molecules  of  the  blood-serum 
are  electrolytes,  and  about  three-fourths  of  these  an*  molecules  of  NaCl.  most 
of  which  are  in  the  dissociated  states 

1  The  current  statement  that  corpuscles  are  impermeable  for  calcium  is  refuted 
by  Ilamlnirgcr   (Zeit.  physikal.  ("hem..  1000   (CO),  C0.3). 
"  2  Ayiiaud.  Ann.  Inst.  Pasteur,  1011    (25),  56. 

2a  Literature  given  by  Powe.  Arch.  Int.  j\Ied..  lOlfi    (IS),  455. 

3  Concerning  relation  of  conductivil v  to  free/.ing-poiiif  see  Wilson.  .Xmer.  .Tour, 
of  Physiol.,  1006   (16),  438. 


THE  COMI'OHITIOX  OF  THE  BLOOD  291 

Enzymes. — A  large  number  of  enzymes  exist  in  tlie  blood,  the  followiufi;  being 
among  those  tliat  have  been  detected:  diastase,  glucase,  lipase,  thrombin,  rennin, 
and  proteases.  The  proteases  and  [lerhaps  the  other  enzymes  are  held  in  elieck 
to  a  large  extent  by  ''antifer7HCtits''  that  are  also  present  (see  "Hnzyines'") .  In 
relation  to  the  antiferments  are  the  innumerable  antibodies  tiiat  exist  normally 
in  the  serum  for  foreign  proteins,  foreign  cells,  and  for  bacteria  and  their  toxins, 
as  well  as  those  resulting  from  reaction  to  infection,  etc. 

The  proportions  in  which  the  constituents  of  the  plasma  normally  occur  have 
been  determined  by  Hoppe-Seyler  and  by  Ilammarsten  as  follows:  * 

Table  I 

No.  1  Xo.  2 

Water 908.4  1)17.6 

Solids         91.6  S2.4 

Total  proteins 77.6  69.5 

Fibrin 10.1  6.5 

Globulin 38.4 

Seralbumin 24.6 

Fat 1.2 

Extractive   substances 4.0  ,„„ 

^  ^             Soluble   salts 6.4  ^'^•^ 

"^  Insoluble  salts 1.7 

No.  1  is  an  analysis  by  Hoppe-Seyler. 

No.  2  is  the  average  of  three  analyses  made  by  Hammarsten. 

Reaction. — It  i.s  very  difficult  to  determine  the  exact  reaction  of 
tlie  blood  plasma.  If  we  titrate  with  an  acid,  we  liberate  much  of 
the  alkali  from  the  proteins,  dissociate  all  the  NagCOg  present,  as  well 
as  the  NaHCO.j  and  the  sodium  phosphate,  and  find  in  this  way  that 
the  entire  fresh  blood  contains  neutralizahle  alkali  corresponding  to  a 
solution  of  NajCOg  of  about  0.443  per  cent,  strength  (Strauss).  In 
other  words,  the  blood  has  a  quantity  of  alkali  in  combination  that 
can  be  drawn  upon  to  neutralize  acids  to  the  extent  indicated  by  the 
above  figures.  The  real  alkalinity  of  a  fluid,  however,  is  dependent 
upon  the  number  of  free  OH  ions  in  the  solution ;  and  Hober  has  de- 
termined by  physico-chemical  methods  that  the  concentration  of  OH 
ions  in  blood  is  but  little  greater  than  in  distilled  water."  Michaelis  ^* 
has  found  the  H+  concentration  of  the  blood  to  be  0.45  X  10"^,  as 
contrasted  with  neutrality  at  38°  which  is  H+  =  1.5  X  10"^  The  in- 
terchange between  COo,  phosphates  and  carbonates  in  the  blood  is 
such  that  it  is  impossible  for  any  considerable  quantities  of  free  H  or 
OH  ions  to  exist,  and  the  protoplasm  is  thus  protected  from  an  excess 
of  either.  The  capacity  of  the  blood  to  neutralize  acids  and  alkalies 
is  sometimes  referred  to  as  its  "buffer  value."  ^^  According  to  Hen- 
derson ^  not  more  than  five  parts  of  excess  free  H  or  OH  ions  can  be 
present  in  ten  billion  parts  of  protoplasm.  An  alkalinity  is  impos- 
sible because  this  would  cause  an  increased  osmotic  pressure  which 

4  For  complete  analyses  of  the  blood  see  Abderhalden,  Zeit.  physiol.  Chem., 
1898   (25),  106. 

^>  For  bibliograpbv  on  Alkalinity  of  Blood,  see  Henderson,  Ergebnisse  Physiol., 
1909   (8),  254. 

saDeut.  med.  Woch..  1914   (40).  1170. 

5b  See  'Levx  and  Rowntree.  Arch.  Int.  Med.,  1916  (17),  525. 

6  Amer.  Jour.  Phvsiol.,  1907   (18),  250;  1908   (21),  427. 


292  DISTURBANCES    OF    CIRCULATIOX 

the  kidneys  would  reflate ;  acidity  is  impossible  because  death  would 
result  from  the  inability  of  the  blood  to  carry  COj.  The  blood  and 
tissue  proteins  also  can  bind  much  of  either  H  or  OH  ions/  so  that  the 
preservation  of  neutrality  is  elaborately  guarded.  In  the  tissues,  be- 
cause of  the  production  of  acids  during  metabolism,  the  H-ion  con- 
centration is  slightly  higher  than  that  of  the  blood,  being  estimated  by 
JVIichaelis  at  exact  neutrality,  1.5  X  10"'^.  Presumably  one  important 
purpose  of  the  exact  regulation  of  reaction  is  to  provide  proper  con- 
ditions for  enzyme  action. 

The  alkali  of  the  blood  exists  in  part  as  alkaline  salts,  carbonate 
and  phosphate  (the  diffusible  alkali),  and  partly  combined  with  pro- 
tein {non-diffusible  alkali).  As  the  corpuscles  are  richer  in  diffusible 
alkali  than  the  plasma  or  serum,  the  number  of  corpuscles  modifies 
the  alkalinity  of  the  blood  decidedly.  j\Iuch  importance  is  attached 
to  the  question  of  the  alkalinity  of  the  blood  for  two  reasons :  first,  in 
certain  conditions  of  disease  the  blood  contains  so  much  of  organic 
acids  that  the  alkali  is  partly  saturated  and  the  power  of  the  blood 
to  carry  CO,  is  lessened,  with  serious  results  (see  "Acid  Intoxica- 
tion," Chap,  xviii)  ;  and,  second,  the  bactericidal  power  of  the  blood 
is  found  to  vary  according  to  its  alkalinity.  In  fact,  metabolic  activ- 
ity seems  generally  to  be  favored  by  certain  degrees  of  alkalinity ;  for 
example,  J.  Loeb  ^  found  that  sea-urchin  eggs  develop  with  much 
greater  rapidity  if  a  small  amount  of  OH  ions  is  free  in  the  sea- 
water.  Brandenburg  ^  states  that  the  non-diffusible  alkali  varies  ac- 
cording to  the  amount  of  protein  in  the  blood ;  in  pneumonia  and 
acute  nephritis  he  found  it  low.  In  cancer  the  titrable  alkalinity  is 
distinctly  increased,  and  IMoore  and  Walker  ^°  find  that  this  is  due  to 
an  increased  alkalinity  of  the  proteins  of  the  blood.  Orlowsky  ^- 
could  find  no  decrease  in  the  alkalinity  of  the  blood  in  leucoeytosis, 
or  when  virulent,  bacteria  were  introduced  into  the  blood.  By  gas 
chain  measurements  of  H  and  OH  concentration,  Roily  '^^  found  prac- 
tically no  change  in  reaction  during  starvation  or  after  bilateral 
nephrectomy.  Abnormally  low  alkalescence  was  rarely  found  except 
in  diabetic  acidosis,  while  increased  alkalescence  was  obtained  chiefly  in 
liver  diseases.  Awerbach  ^^  claims  that  in  severe  high  fevers  the  bac- 
tericidal effect  of  the  blood  alkalinity  is  increased  (see  also  "Passive 
Congestion"  for  further  discussion  concerning  the  relation  of  alka- 
linity to  bactericidal  power). 

Viscosity  of  the  Blood.^" — Xonnal  l)lood  is  about  five  times   (4.5 

7  See  Robertson.  Jour.  Uiol.  Cliem.,  1909   (fi),  313;   1910   (7).  3.">1. 

8  Arch.  f.  Ent.\vicklun<:sinoolianik,   189S    (7),  031. 

oDout.  mod.  Woch.,  1902   (28),  78;  Zeit.  f.  klin.  ^fed..  1902   (45),  l.>7. 
If*  Biocliem.  Jour.,   1900    (1),  297;   pood  discussion  of  blood  reaction. 
12  Dent.  mod.  Wocli.,   1903    (29).  (iOl. 
i3Miincb.  nied.  Wocli..  1912    (r)9),  1201  and  1274. 

14  Med.  Obosrenijo,   1903,  p.  .')90. 

15  Review  of  literature  by  Determann.   Zeit.  klin.  Med.,   1910    (70),   185;    also 


HEMORRHAGE  293 

times,  Austrian)  more  viscous  than  water,  chiefly  because  of  the 
corpuscles  and  the  dissolved  proteins.  This  viscosity  does  not  vary 
directly  Avith  the  specific  gravity  or  the  hemoglobin,  but  is  closely 
related  to  the  number  of  red  corpuscles  (Burton-Opitz)  ;  laking  the 
corpuscles  increases  the  viscosity  considerably.  ^lost  salts  increase 
the  viseosit}',  but  some,  especially  iodides,  are  said  to  reduce  it.  Car- 
bon dioxide  increases  viscosity  greatly,  even  when  in  amounts  possible 
in  the  circulating  blood.  Anemia  decreases  the  viscosity,  approxi- 
mateh'  in  proportion  to  the  number  of  corpuscles ;  polycythemia  is  ac- 
companied by  a  corresponding  increase ;  leukemia,  because  of  anemia, 
shows  a  decrease ;  in  nephritis  there  may  either  be  an  increase  or  a  de- 
crease in  the  viscosity,  not  corresponding  in  any  way  to  the  blood  pres- 
sure. Cardiac  disease  with  edema  shows  low  viscosity"  because  of  the 
anemia  and  hydremia,  but  if  there  is  polycythemia  and  no  edema  the 
viscosity  may  be  high.  Jaundice,  causes  an  increase,  diabetes  gives 
variable  results.  Typhoid  causes  no  characteristic  change  beyond  that 
resulting  from  anemia,  and  in  pneumonia  the  cyanosis  and  salt  reten- 
tion usually  cause  an  increase  (Austrian).  Gullbriug  ^^'^  found  the 
viscosity  to  vary  directl}'  with  the  per  cent,  of  neutrophiles. 

HEMORRHAGE 

Hemorrhages  result  from  an  altered  condition  in  the  vessel-walls, 
which  may  be  due  either  to  trauma  or  to  chemical  injuries.  Of 
the  chemical  agencies  causing  hemorrhages,  bacterial  products  are  the 
most  important  practically,  but  manj'  poisons,  such  as  phosphoinis, 
formaldehyde,  phytotoxins  (ricin,  abrin,  and  crotin),  and  zootoxins 
(snake  venoms)  cause  numerous  and  abundant  hemorrhages.  For- 
merh',  the  tendency  was  to  ascribe  hemorrhages  from  the  above  causes 
to  mechanical  injury  of  the  vessels  by  thrombi,  or  by  emboli  of  ag- 
glutinated corpuscles,  but  the  work  of  Flexner "  has  shown  that 
venoms  cause  hemorrhages  by  injuring  the  capillary  walls,  so  that 
actual  rents  are  produced  by  the  intravascular  pressure,  and  it  seems 
highly  probable  that  hemorrhages  are  produced  by  other  chemical 
substances  in  a  similar  way.  We  may,  therefore,  refer  such  hemor- 
rhages to  an  endotheliotoxic  action  of  the  poison,  or  to  a  solvent  effect 
upon  the  intercellular  cement  substance.  In  the  case  of  ordinary 
chemical  poisons  the  endotheliotoxic  action  is  not  specific,  but  with 
some  of  the  toxins  it  seems  to  be  quite  so;  for  example,  rattlesnake 
venom  contains  an  endotheliotoxic  substance  (hemorrhagin) ,  which 
seems  to  be  a  specific  poison  for  endothelium,  and  which  is  the  most 
dangerous  constituent  of  the  venom.  If  we  immunize  animals  against 
tissues  containing  iiiucli  endotlielium  (  c.  g.,  lymph-glands),  their  serum 

Austrian,  Johns  Hopkins  Hosp.  Bull.,   1911    (22),  9.     See  also  Traube,  Internat 
Zeit.  phvsik.-chem.  Biol.,   1014    (1),  380. 

isaBeitr.  klin.  Tuberk.,  1014    (30),  1. 

16  Univ.  of  Penn.  Med.  Bull.,  1902   (15),  355. 


294  DISTURBANCES    OF    CIRCULATION 

will  be  found  to  contain  endotlieliotoxins,  so  that  when  this  serum 
is  injected  subcutaneously  into  a  susceptible  animal,  large  local  hemor- 
rhages result;  if  injected  into  the  peritoneal  cavity,  there  results 
marked  desquamation  of  the  endotlielial  cells,  which  soon  undergo  de- 
generative changes  (Ricketts).^^  It  is  quite  probable  that  the  bac- 
terial poisons  that  cause  marked  hemorrhagic  manifestations  likewise 
contain  endotlieliotoxins,  although  tliis  matter  does  not  seem  to  have 
been  investigated. 

Even  hemorrhage  by  diapcdesis  seems  to  be  due  to,  or  at  least 
associated  with,  chemical  changes  in  the  capillary  walls,  for  Arnold  ^* 
found  that  when  capillaries  from  M^hich  diapedesis  had  occurred 
were  stained  by  silver  nitrate,  dark  areas  were  found  between  the 
endothelial  cells.  As  silver  nitrate  is  a  stain  for  chlorides,  and  dark- 
ens intercellular  substance  because  it  is  rich  in  sodium  chloride 
(Macallum),  it  is  probable  that  there  is  an  increase  in  the  amount 
or  a  difference  in  the  method  of  combination  of  the  chlorides  of  the 
cement  substance  between  the  endothelial  cells  at  the  places  where 
red  corpuscles  escape.  ]\I.  H.  Fischer  ^^  suggests  that  diapedesis  re- 
sults from  a  change  in  the  endothelial  cells,  which  under  the  influence 
of  acids  or  other  agents  of  metabolic  origin  become  excessively  hydro- 
philic,  swell  up,  and  become  so  softened  that  corpuscles  may  pass  di- 
rectly through  the  cell,  just  as  a  drop  of  mercury  can  pass  through 
a  sufficiently  soft  jelly  without  leaving  a  hole  in  the  jelly. 

Hemorrhage  in  cachetic  conditions  is  often  ascribed  to  changes 
in  the  vessel-walls  due  to  malnutrition,  but  it  is  difficult  to  imagine 
capillary  walls  suffering  from  lack  of  nourishment,  even  with  the 
poorest  of  blood,  and  it  seems  more  probable  that  the  hemorrhages  are 
due,  even  in  cachexia,  to  chemical  constituents  of  the  blood  that  in- 
jure the  endothelium.  Hemorrhages  that  follow  re-establishment  of 
the  circulation  after  complete  occlusion,  however,  may  be  the  result 
of  asphyxial  changes  in  the  capillary  walls,  presumably  colloidal  swell- 
ing of  the  cells. 

After  severe  hemorrhages  the  blood  shows  a  decrease  in  specific 
gravity  and  viscosity,  an  increase  in  surface  tension  and  electrical 
resistance,  and  either  increase  or  decrease  of  the  freezing-point  de- 
pression, all  these  changes  being  transient  if  the  individual  is  other- 
wise normal.-"  (See  also  Secondary  Anemia.)  There  is  a  rapid 
absorption  of  fluid  from  the  tissues  and  tissue  spaces,  resulting  in  a 
dilution  of  protein  and  formed  elements,  but  not  of  salts.  There  is 
?aid  to  be  a  decreased  permeability  of  vessels,  resulting  in  reduced 
exudative  processes.^ "^     The  proportion  of  the  several  blood  proteins 

17  Trans.  Chicago  Path.  Soc,  1902    (5),  181. 

18  Virehow's  Arch.,  1S75    (62),  157. 

m  "Nophriiis,"  New  ^'ork,   1012,  p.  78. 
2"01iva,  Folia  cliiiicii,  1!»12    C?),  21:^ 
»«ii.Liiitlil('ii,  M.'d.   Klin..  1913    (9),  1713. 


iiKMoiiiiiiAdi:  295 

is  variably  altci't'd  aftrr  repeated  lieiuorrliag'es;  the  sugar  is  little 
affected  l>ut  the  non-jjroteiu  nitrogen  and  urea  are  increased.'''"  Rapid 
lieniorrhagos  cause  a  decrease  in  the  coagulation  time  l)ecause  of  a 
decrease  in  antitlirond)in  and  a  sligiit  increase  in  prothrombin,  in  spite 
of  a  decrease  in  fibrinogen.^-"  If  the  blood  is  withdrawn  repeatedly 
in  large  amounts,  centrifuged,  and  the  washed  corpuscles  reinjected 
suspended  in  isotonic  salt  solution  (plasmaphaeresis),  life  can  be 
maintained  even  after  4  to  5  times  the  total  volume  of  blood  has  l)een 
removed  and  washed.  This  is  possible  because  of  rapid  reformation 
of  the  plasma,  and  the  ])lood  sliows  the  changes  characteristic  of  sec- 
ondary anemias. ^"'^ 

Changes  in  the  Extravasated  Blood. — These  begin  soon  after 
its  escape.  In  most  situations  sufficient  fibrin  ferment  is  formed  to 
cause  prompt  clotting,  but  in  the  pleura  and  other  serous  cavities  the 
blood  may  remain  fluid  for  some  time,  possibly  because  of  lack  of 
cellular  injury  that  miglit  cause  liberation  of  fibrin  ferment.-''^  If  the 
blood  does  not  become  infected,  the  rapidity,  of  subsequent  changes 
depends  chiefly  upon  the  location  and  amount  of  blood.  Small  ex- 
travasations of  blood  into  the  tissues  are  subjected  to  the  action  of  the 
tissue  cells  and  of  leucocytes  emigrating  freely  from  the  capillaries; 
large  masses  of  blood  are  but  little  affected  by  these  agencies,  the 
leucocytes  within  the  mass  soon  die,  and  secondary  changes  go  on  very 
slowly.  In  small  subcutaneous  hemorrhages  (e.  (/.,  a  bruise)  enzymes 
from  the  invading  leucocytes  and  tissue-cells  soon  dissolve  the  small 
quantities  of  fibrin  present ;  even  earlier  the  stroma  of  the  red  cor- 
puscles is  so  altered  that  hemolysis  occurs  and  the  hemoglobin  escapes 
and  diffuses  into  the  tissues.  This  hemolysis  may  be  brought  about 
by  the  action  of  proteolytic  enzjTnes  on  the  corpuscles,  or  by  the  hemo- 
lytic action  of  the  products  of  protein  splitting.  Soon  the  hemoglo- 
bin disintegrates,  forming  the  masses  of  pigment  so  characteristic 
of  old  hemorrhagic  areas,  and  also  giving  rise  to  the  discoloration 
observed  beneath  the  skin  in  the  later  stages  of  resolution  of  hemor- 
rhagic extravasations.  The  first  products  of  the  splitting  of  hemo- 
globin are:  (1)  The  protein,  glohin,  which  constitutes  94  per  cent, 
of  the  hemoglobin;  and  (2)  the  iron-containing  coloring-matter,  hem- 
atin  (in  the  absence  of  oxygen  the  pigment  is  reduced  hematin  or 
hemochromogen).  As  hematin  may  be  experimentally  obtained  by 
the  action  of  proteases  upon  hemoglobin,  the  decomposition  of  the 
hemoglobin  in  the  tissues  is  probably  accomplished  in  a  similar  way 

lOb  Taylor  and  Lewis,  Jour.  'Riol.  Chom.,  19X5   ^22),  71. 

i9i- Drinker,  Amer.  .Jour.  Plivsiol.,  1915   '.36).  :^0o. 

iMAbel  et  al.  .Tour,  rharmacol.,  Ifll4   (5).  (i2.5 ;  101,5   (7),  120. 

20a  Denny  and  Minot  (Amor.  Jour.  Physiol.,  1016  (.30).  4-55)  believe  that  the 
blood  really  does  clot,  and  that  it  remains  lluid  when  withdrawn  because  the 
fibrinogen  has  been  removed  by  olottino;.  Zahn  and  Walker  (Biochem.  Zeit..  101.3 
(58).  130).  however,  consider  that  the  fibrinogen  is  altered  by  the  pleural  endo- 
thelium, so  that  it  cannot  clot. 


296  DISTURBAXCES    OF    CIRCULATION 

b}^  the  proteases  of  the  leueoeytes,  tissue-cells  and  blood  plasma ;  the 
g'lobin  is  thus  digested  away  and  the  soluble  products  carried  off, 
while  the  insoluble  liematin  remains.-^  The  hematin  gradually  un- 
dergoes further  changes,  forming  an  iron-free  i)igment  (hcinatoulin) 
and  an  iron-containing  pigment  (hemosiderin) . 

Hematoidin  is  nearly  or  quite  identical  with  the  bile-pigment,  hili- 
rubin,  and  is  absorbed  from  the  hemorrhagic  extravasation  and  elimi- 
nated as  bilirubin  in  the  bile.  Possibly  some  of  the  hematoidin  un- 
dergoes transformation  into  nrobiUn,  and  is  then  eliminated  in  the 
urine.  Hemosiderin  seems  to  be  relatively  insoluble  and,  therefore, 
is  more  slowly  removed,  so  that  it  may  be  found  at  the  site  of  a  hem- 
orrhage after  the  other  evidences  of  blood  extravasation  have  been 
removed.  It  may  be  easily  demonstrated  by  staining  with  potassium 
ferrocyanide,  the  Prussian  blue  that  is  formed  being  readily  dis- 
tinguished. Unstained  hemosiderin  generally  appears  in  the  form 
of  brown  or  yellowish-brown  granules,  never  as  crystals.  After  a 
time  the  hemosiderin  is  taken  away,  and  probably  is  to  a  greater  or 
less  extent  deposited  in  the  liver  and  spleen,  either  as  hemosiderin 
or  as  some  other  iron  compound.  Eventually  it  is  probably  utilized 
to  make  new  hemoglobin  ;  at  any  rate,  the  iron  liberated  by  the  breaking- 
up  of  hematin  within  the  body  does  not  appear  to  be  eliminated. "- 

The  changes  in  the  red  corpuscles  described  above  are  not  at  all 
peculiar  to  extravasated  blood,  but  are  quite  the  same  as  the  changes 
that  are  going  on  continuously  and  normally  in  the  blood.  Red  cor- 
puscles are  short-lived,  being  but  non-nucleated  fragments  of  cells, 
and  they  are  continually  disintegrating  with  the  production  of  iron- 
free  pigments  that  are  excreted  as  the  coloring-matters  of  the  bile  and 
urine,  while  the  iron  is  worked  over  again  into  new  hemoglobin 
after  a  varying  period  of  storage  in  the  tissues,  particularly  in  the 
spleen  and  liver.  The  destruction  of  red  corpuscles  under  normal 
conditions  seems  to  take  place  chiefly  in  the  spleen,  bone-marrow, 
and  hemolymph  glands,  where  injured  or  decrepit  c()ri)uscles  are 
taken  out  of  the  blood  by  the  phagocytic  endothelial  cells,  and  de- 
composed by  intracellular  enzymes.  In  hemorrhagic  extravasations 
the  changes  are  essentially  the  same;  some  corpuscles  are  destroyed 
by  phagocytes,  but  more  by  extracellular  enzymes.  The  products 
of  decomposition  also  seem  to  be  no  different  from  those  formed  dur- 
ing normal  katabolism  of  hemoglobin,  and  they  meet  the  same  fate  in 
the  end. 

If  the  hemorrhages  are  very  abundant,  some  hemoglobin  may  be 
absorbed  as  such  and  appear  in  the  urine,  but  this  ])rol)nbly  seldom 
happens  unless  red  corpus('le^'.  are  also  being  destroyed  in  tlie  circu- 
lating  blood.--''     An   increased   amount   of   iron    accumulates   in    the 

21  More  fullv  disciisscHl  in  tlie  consideration  of  "Pitrnu'ntaiion."  Clia|).  xvi. 

22  See  Morishima,  Arch.  f.  cxp.  Tath.,  1898    (41),  2!)]. 

22a  In   cerebral   lieniorrliase   the  blood    serum    niav   be   frreenisli    and    somewhat 


IIEMOI'IIILIA  297 

liver,  but  if  iniicli  blood  has  been  lost  by  lieiiioiThage  on  free  surfaces, 
the  iron  conteiit  of  the  liver  is  decreased,  as  it  is  taken  away  to  form 
new  hemoglobin  (Quincke).-^  Excretion  of  bile-pigments  is  in- 
creased by  destruction  of  blood  (Stadelmann),  but  not  greatly  in 
the  case  of  hemorrhages,  for  the  blood  is  decomposed  and  absorbed 
too  slowly.  Schurig  -*  found  that  hemoglobin  injected  into  the  tis- 
sues is  partly  decomi)osed  in  situ  with  formation  of  iron  compounds, 
but  the  greater  part  enters  the  circulation  as  hemoglobin,  and  is 
partly  converted  into  bile-pigment  by  the  liver-cells,  the  rest  being 
converted  into  simpler  iron  compounds  by  the  spleen,  bone-marrow, 
and  renal  cortex. 

If  the  hemorrhagic  extravasation  has  been  large  in  amount,  the 
deeper  ])ortioiis  of  the  mass  are  not  soon,  if  ever,  invaded  by  leuco- 
cytes or  tissue-cells.  Consequently  the  blood  is  acted  upon  very 
slowly  by  the  enzymes  liberated  by  the  leucocytes  it  contains  itself, 
and  by  the  small  amounts  of  proteases  in  the  serum.  Furthermore, 
the  products  of  decomposition  are  not  soon  absorbed,  but  accumulate 
in  considerable  amounts,  so  that  we  often  find  crystalline  deposits  of 
hematoidin,  sometimes  even  of  hematin,  hemoglobin,  or  parahcmoglo- 
hin-  (Nencki)  -"'  or  methemogJohin. 

The  least  soluble  constituent  of  the  red  corpuscle  stroma,  choles- 
terol, also  accumulates  in  such  extravasations  as  large,  thin  plates ; 
after  most  of- the  other  products  of  distintegration  have  been  absorbed 
from  such  accumulations  of  blood,  the  most  conspicuous  part  of  the 
residue  may  be  a  mass  of  cholesterol  crystals  imbedded  in  prolifer- 
ating connective  tissue. 

HEMOPHILIA  2G 

There  are  several  pathological  conditions  associated  with  increased 
tendency  to  bleeding,  notably  scurvy  and  the  various  forms  of  pur- 
puras, but  especially  the  remarkable  hereditary  condition,  hemophilia. 
In  the  purpuric  diseases  various  of  the  factors  concerned  in  coagula- 
tion of  the  blood  have  been  found  altered,-"'''  notably  the  blood  plate- 
lets,-^'' but  Howell  found  no  change  in  either  prothrombin  or  anti- 
thrombin  in  purpura  hemorrhagica  and  other  related  conditions. 
Similar  negative  results  were  obtained  in  scurvy  by  Hess.-*"=     Melena 

fluorescent  from  absorbed  pigment,  according  to  !Marie  and  Lcri,  Union  Pharm., 
Aug.  15,   1914. 

23Deut.  Arch.  klin.  Med.,  1880   (25),  567:   1880   (27),  103.  , 

24  Arch.  exp.  Path.  u.  Pharm.,   1808    (41),  29. 

23  Arch.  exp.  Path.  u.  Pharm.,  1886   (20),  .3.S2. 

26  Literature  and  resume  given  bv  Stonipcl,  Cent.  f.  Grenzgel).  ^Ted.  ni.  Chir.. 
1900  (3).  753;  Sahli,  Zeit.  f.  klin.  :\ied..  lOO.i  (56),  294:  :Marchand,  in  Krehl  and 
Marchand's  Handb.  allg.  Pathol.,  1912.  II  (1),  307.  Also  later  references  in  this 
text. 

26a  See  Hurwitz  and  Lucas,  Arch.  Int.  Med..  1916  (17),  543:  Minot  ei  ah,  ibid.^ 
1916  (17),  101.  r">. 

26b  See  Lee  and  Robertson,  Jour.  Med.  Res.,  1916  (33>,  32*r 

26cAmer.  Jour.  Dis.  Children,  1914   (8),  386. 


298  iJlSTURBAyvE.S    OF    CIRCULATION 

neonatorum  exhibits  decreased  prothrombiu  in  tlie  blood,  while  in 
leukemias  and  anemias  there  may  be  an  excess  of  antithrombin,-"'' 
leading  to  severe  hemorrhage  (see  also  Thrombosis). 

Since  hemophilia  seems,  superficially  at  least,  to  depend  upon  some 
alteration  in  a  chemical  property  of  the  blood,  namely,  coagulability, 
it  is  frequently  i-egarded  as  an  example  of  hereditary  transmission 
of  a  chemical  abnormality.  The  exact  cause  of  this  peculiar  tendency 
to  prolonged  bleeding  from  insignificant  or  perhaps  imperceptible 
wounds  has  been  sought  vigorously  by  both  histological  and  chemical 
means,  but  as  yet  without  avail.  Various  observers  have  described 
abnormal  thinness,  or  increased  cellularity  or  fatty  degeneration  of 
the  vessel-walls,  but  the  findings  have  been  far  too  inconstant  to 
afford  a  satisfactory  anatomical  explanation  of  all  the  features  of 
hemophilia.  Likewise  increased  blood  pressure  can  be  ruled  out,  for 
although  the  left  heart  is  frequently  enlarged,  there  is  usually  no  in- 
creased blood  pressure  demonstrable;  furthermore,  conditions  of 
high  blood  pressure,  such  as  nephritis,  do  not  cause  hemophilia.  The 
theory  of  "hydremic  plethora"  is  also  without  good  foundation. 

The  most  natural  place  to  look  for  the  fundamental  fault  is  in  the 
blood,  but  speaking  strongly  against  this  is  the  occasional  occurrence 
of  "local"  hemophilia;  e.  g.,  in  this  type  of  hemophilia  wounds  of  the 
skin  may  behave  as  in  normal  individuals,  whereas  any  injury  of  the 
mucous  surfaces  is  followed  by  pronounced  hemophilic  bleeding ;  - ' 
in  other  cases  the  hemophilic  bleeding  is  limited  to  regions  above  the 
shoulders;  in  still  another  class  the  bleeding  is  always  from  one  or- 
gan, e.  g.,  the  kidney's.  Nevertheless,  a  great  deal  of  investigation  of 
the  blood  has  been  done,  at  first  chiefly  with  negative  results.  There 
are  no  characteristic  changes  in  the  cellular  elements  of  the  blood, 
beyond  the  changes  common  to  all  secondary  anemias,  excepting  pos- 
sibly a  decrease  in  the  number  of  white  corpuscles  with  a  relative 
increase  in  the  number  of  lymphocytes  as  observed  hy  Sahli;  the 
platelet  count  is  normal.  No  constant  alterations  in  the  salts  of  the 
blood  have  been  found,  calcium  usually  being  normal ;  and  the  propor- 
tion of  water,  fibrinogen  and  the  several  other  proteins,  the  alkalinity, 
and  the  osmotic  pressure  of  the  serum  all  seem  to  be  normal.  IMetab- 
olism  is  unchanged,  except  possibly  for  calcium  loss  in  some  cases.-* 
Since  bleeding  is  normally  stopped  principally  by  coagulation,  a  de- 
ficiency in  fibrin  oi'  its  antecedents  might  be  expected,  but  most 
studies  on  this  point  have  shown  a  noi-mal  amount  of  fibrinogen  in 
the  blood  of  hemophilics,  the  fre(pient  formation  of  large  tumors  of 
clotted  blood  at  the  bleeding  points  supporting  the  experimental 
evidence  that  the  blood  contains  an  abundance  of  fibrinogen.     The 

2od  Whipple,  Arcli.  Int.  Aled.,  1913   (12),  637. 
27  Ahdi'ilialdcii,   Zicfrl,.,'s   I'.oiir.,   1!)04    ( .T) ) .  '213. 

2«  Kulin,  Amer.  Jour.  Dis.  Cliildroii,  lOKJ  (  11 ),  103:  Laws  and  Cowie,  ibid.,  1917 
(13),  236;  Hess,  Bull.  Joiins  Hopkins  Hosp.,  1916   (26),  372. 


iiKMoi'iiii.ix  299 

■'blectliii^'  time""  t'ullowing  puiiclurcs  in  tliu  skin  is  not  excessive.  As 
to  the  rate  of  clotting,  Sahli,-°  who  avoided  a  number  of  errors  made 
in  earlier  investio-ations,  found  that  in  the  intervals  between  the  at- 
tacks of  hemorrhafi-c  the  rate  of  the  coa^'ulation  of  the  blood  is  con- 
stantly Huicli  slower  than  noi-iiial.  Dui'iii;^-  an  attack  of  bleeding  the 
coagulation  time  approaches  the  normal ;  indeed,  it  may  be  faster 
than  normal ;  apparently  this  is  due  to  a  reaction  on  the  part  of  the 
organism  to  the  loss  of  blood.  If  blood  is  collected  directly  from  the 
site  of  bleeding  the  coagulation  time  is  very  rapid,  because  of  the  ac- 
cumulation of  fibrin  ferment  from  the  clot  over  which  the  escaping 
blood  flows.  Yet  in  spite  of  the  normal  coagidability  of  the  blood  and 
the  rapid  clotting  after  the  blood  escapes  from  the  vessel,  bleeding 
continues  for  long  periods  before  it  can  be  stopped.  As  he  found  no 
general  change  in  the  properties  of  the  blood  to  account  for  the 
bleeding,  and  as  local  influences  seem  to  be  important  in  hemophilia, 
Sahli  advanced  the  plausible  hypothesis  that  the  cause  of  hemophilia 
lies  in  hereditary  deficiency  of  the  fibrin-forming  substances,  throm- 
bokinase  or  zymoplastic  substance  (see  "Thrombosis"),  in  the  vessel- 
walls,  so  that  when  the  vessels  are  injured  there  is  no  local  produc- 
tion of  fibrin  such  as  occurs  normally.  Local  hemo])hilia  may  be  ex- 
plained readily  as  a  local  deficiency  in  fibrinoplastic  material.  In 
general  hemophilia  even  the  leucocytes  may  exhibit  the  same  defect, 
in  which  case  clotting  of  the  blood  is  diminished  even  outside  the  tis- 
sues. This  hypothesis  seems  to  be  in  excellent  agreement  with  many 
of  the  facts  now  known,  but  there  yet  remains  to  be  demonstrated 
such  a  lack  of  fibrin-forming  elements  in  the  vessel-walls  and  other 
tissues  of  a  hemophilic  subject,  and  a  single  autopsy  of  a  hemolytic 
subject  gave,  on  the  contrarj^  a  very  active  thromboplastic  extract  from 
the  vessels  fGressot).-^ 

"With  the  improved  methods  of  study  of  the  factors  in  coagulation 
of  blood  introduced  by  Howell,  it  has  been  found  by  him  and  cor- 
roborated by  others  ^°  that  in  hemophilia  there  is  constantly  a  defi- 
ciency in  prothrombin,  the  other  factors  being  practically  normal  in 
amount,  and  as  in  other  hemorrhagic  conditions  there  is  no  equal 
alteration  in  the  prothrombin,  they  look  upon  this  change  as  an 
essential  characteristic  of  hemophilia.  Fonio,  and  ]\Iinot  and  Lee, 
however,  find  that  the  blond  platelets  of  hemophilics  are  remarkably 
ineffective  in  causing  coagulation  of  either  normal  or  hemophilic 
plasma,  although  normal  platelets  cause  normal  coagulation  of  hemo- 
philic plasma,  and  therefore  conclude  that  there  is  some  deficient  ac- 
tivity on  the  part  of  the  platelets  in  spite  of  their  occurrence  in  normal 
luimbers  in  hemophilia.     The  significance  of  the  platelets  is  shown 

20  Zeit.  klin.  Med.,  1012    (70),  104.     Since  corroborated  by  Minot  and  Lee. 

30  Howell,  Arch.  Int.  Med.,  1014  (1.3),  76;  Hurwitz  and  Lucas.  ihicL.  1016  (17), 
543:  ^linot  and  Lee,  ihid.,  1010  (18),  474;  these  papers  review  re<'ent  work  on 
hemophilia. 


300  DLSTCRliAXCIJS    OF    CIRCCLATION 

especiallj"  clearly  by  the  observation  of  Ledingham  and  Bedson  ^^ 
that  anti])lat('let  seruiu  will  ])rotliice  a  ]Mir]nirie  condition  when 
injected  into  animals  of  the  species  furnishing  the  platelets,  al- 
though no  similar  effect  is  produced  by  antileucocyte  or  antiery- 
throcyte  serum.  Hess^-  states  that  there  may  be  an  hereditary 
jiurpura,  sometimes  occurring  in  the  females  of  hemophilic  families, 
differing  from  hemophilia  in  a  deficiency  in  the  number  of  platelets, 
hemorrhages  following  local  congestions  or  puncture  wounds  and 
exhibiting  an  increase  in  the  bleeding  time. 

ANEMIA  AND  THE  SPECIFIC  ANEMIAS  ^^ 

The  customar}^  division  of  the  anemias  is  into — (a)  priinary,  i.  e., 
those  in  which  the  anemia  seems  to  depend  upon  some  abnormality  in 
the  blood-forming  organs  or  in  the  blood  itself;  and  (&)  secondary, 
embracing  anemias  the  result  of  some  obvious  cause,  such  as  hemor- 
rhage, poisoning  with  blood-destroying  poisons,  cachexia,  etc.  In 
these  various  forms  of  anemia  certain  chemical  differences  prevail,  but 
they  are  by  no  means  so  striking  as  are  the  histological  differences  in 
the  formed  elements  of  the  blood. ^* 

SECONDARY  ANEMIAS 

As  the  simplest  variety,  anemia  following  a  single  large  hemorrhage 
may  be  considered  first. 

If  loss  of  blood  by  hemorrhage  is  rapid,  the  effects  are  naturally 
much  more  serious  than  when  the  loss  is  slow.  The  total  quantity'  of 
blood  in  the  average  adult  is  estimated  at  about  ]{r,  to  ^i.j  the  total 
body  weight  (therefore  about  10  to  12  pounds),  although  this  pro- 
portion does  not  hold  for  extremely  obese  or  extremely  thin  indi- 
viduals; ^^  in  infants  the  proportion  is  lower — about  ^20-  When  one- 
third  of  the  total  amount  of  blood  is  lost  rapidly,  a  marked  fall  of 
blood  pressure  occurs;  loss  of  one-half  of  the  total  amount  may  be 
fatal,  and  loss  of  more  than  that  at  one  time  usually  is  fatal.  The 
chief  cause  of  death  following  large  hemorrhages  is  the  low  blood 
pressure  rather  than  the  loss  of  any  of  the  constituents  of  the  blood ; 
hence  the  successful  results  of  the  use  of  physiological  salt  solution 
after  severe  hemorrhage.  The  number  of  corpuscles  may  be  greatl.v 
reduced  after  several  small  hemorrhages,  even  to  as  low  as  11  per 
cent,  of  the  normal  number  (Hayem),  without  fatal  results,  because 
in  the  intervals  between  the  hemorrhages  enougli  fluid  lias  been  taken 
up  b}'  the  blood  to  maintain  the  blood  pressuiv  within  safe  limits. 

31  Lancet,  Fcl).  l."?.  101.5. 

32  Arch.  Int.  Med.,  1!)1()    (17),  203. 

33  M(.(abolism  in  anemia  reviewed  I)v   ^lolir,  nandbucli  d.   Bioclioni.,   liUO    ( I\' 
(2)   ),  372. 

34  Concerning  local  anemia,  see  "Infarcts." 

35TIaldane  and  Smith  (Jour,  of  Physiol.,  1000  (25),  331)  estimated  tiie  blood 
of  adults  at  but  J^q  "f  ^'"'  1>o(l.v  weiglit. 


ANEMIA  AND  THE  HPEVIFIC  ANEMIAS  301 

After  a  severe  heiuorrliag-e  tlie  composition  of  the  blood  changes 
rapidly,  for  the  fluids  contained  within  the  tissues  and  lymph-spaces 
nass  into  the  blood  in  large  amounts.  This  helps  to  maintain  blood 
pressure,  but  results  in.  the  blood  containing  a  large  proportion  of 
water  and  salts  and  a  smaller  amount  of  protein  and  red  corpuscles ; 
the  "total  alkalinity"  also  falls,  largely  because  of  the  scarcity  of 
"fixed  alkali,"  on  account  of  the  poverty  in  corpuscles  and  blood 
proteins.  The  proportion  of  water  increases  at  first  more  rapidly 
than  the  proportion  of  salts,  and  as  a  consequence  the  size  of  the  red 
corpuscles  is  increased  because  of  imbibition  of  water ;  indeed,  it  is 
possible  that  this  may  even  be  sufficient  to  cause  hemolysis,  which  wall 
happen  if  the  isotonic  streng-th  of  the  blood  becomes  less  than  that 
of  a  0.46  per  cent.  NaCl  solution  (Limbeck),  while  swelling  may 
occur  whenever  the  strength  is  below  0.8  per  cent.  The  specific 
gravity  of  the  erythrocytes  is  decreased ;  ^^  the  depression  of  the  freez- 
ing point  increases,"  while  the  viscosity  falls.  The  number  of  plate- 
lets is  high. 

Regeneration  of  the  blood  begins  veiy  soon,  and  for  some  tniie  the 
number  of  corpuscles  exceeds  the  proportion  of  hemoglobin.  During 
this  time  the  amount  of  iron  in  the  liver  and  spleen  is  decreased,  it 
being  taken  up  to  be  used  in  the  formation  of  new  hemoglobin.  If 
the  hemorrhages  are  numerous  and  the  condition  of  anemia  prolonged, 
secondary  changes  in  the  viscera  may  occur,  fatty  metamorphosis  be- 
ing most'  marked,  supposedly  because  of  decreased  oxidation.  Indeed, 
many  observers  state  that  repeated  bleedings  greatly  increase  body 
weight  by  causing  increased  fat  deposition. 

Metabolic  Changes. — Gies  ^'^  studied  the  metabolism  of  dogs  after 
Avithdrawing  a  total  amount  of  blood  equal  to  11.5  per  cent,  of  the 
body  weight  during  four  bleedings,  and  found  that  a  slight  and  tem- 
porary increase  in  nitrogenous  elimination  followed  the  bleedings, 
owing  to  an  increased  protein  katabolism.  Sugar  increases  in  the 
blood,  while  albumin  and  laetic  acid  appear  in  the  urine.  After  each 
successive  hemorrhage  the  proportion  of  fibrin  and  the  coagulability 
of  the  blood  increase,  while  the  proportion  of  the  ash  obtained 
from  both  blood  and  serum  remains  practically  unchanged  (:\Ieyer 
and  Gies).  Baumann ''«  states  that  in  regeneration  after  hemorrhage 
the  serum  albumins  increase  more  rapidly  than  the  globulins,  while 
others  have  observed  the  opposite  relation.  The  urine  in  secondary 
anemia  shows  the  effects  of  increased  protein  katabolism.  its  specific 
gravity,  total  solids,  and  total  nitrogen  being  raised ;  the  total  amount 
of  urine  is  at  first  diminished  because  of  lowered  blood  pressure,  but 
it  soon  rises  above  normal  and  later  falls  back  to  normal.     The  view 

soBonninger,  Zeit.  exp.  Path..  1012    (11),  1. 

37  Hoesslin.  Hofnieister's  Boitr..  1906    (8).  431. 

38  American  IMed.,  1004   (8),  155   (resume  of  literature). 

39  Jour.  Phvsiol.,  1903    (29),  18. 


302  DISTURBANCES    OF    CIRCULATION 

formerly  held  tliat  oxidation  is  decreased  in  anemia  has  been  con- 
siderably modified  by  more  recent  investig'aticms;^"  in  fact,  respira- 
tion stndies  indicate  heightened  gas  exchange  in  secondary  anemia.*"* 

Secondary  anemia  due  to  cachexia,  or  to  malnutrition,  is  accom- 
panied ])y  a  general  decrease  in  all  the  elements  of  the  blood,  both 
cellular  and  chemical.  The  proteins  of  the  plasma,  particularly,  show 
a  decrease  in  starvation,  being  drawn  on  by  the  cells  for  food,  and 
the  total  quantity  of  blood  as  well  as  of  each  of  its  constituents  is  de- 
creased (Panum),*^  but  the  proportion  of  blood  to  body  weight  re- 
mains about  normal.  With  protracted  starvation  there  is  only  a 
slight  loss  of  hemoglobin  and  an  increased  coagulability,  but  practi- 
cally no  other  changes.'*^''  In  aplastic  anemias  the  prothrombin  and 
platelet  content  are  likely  to  be  low,  with  normal  fibrinogen. ^^'^ 

Anemia  due  to  hemolytic  agencies  presents  quite  different  fea- 
tures, in  that  red  corpuscles  are  almost  solely  attacked  and  the  prod- 
ucts of  their  disintegration  are  present  in  the  plasma.  As  a  result, 
the  plasma  or  serum  may  contain  free  hemoglobin,  and  if  the  hemo- 
globin is  in  large  amounts,  it  may  escape  into  the  urine.  Thus  par- 
oxysmal hemoglohinuria.  is  probably  due  to  the  presence  in  the  blood 
of  hemolytic  substances,  which  can  be  demonstrated  in  the  blood  of 
the  patients  during  the  attack.  (See  Chapter  viii.)  The  products 
of  the  decomposition  of  the  hemoglobin  set  free  by  hemolysis  are 
present  not  only  in  the  blood,  but  also  in  the  organs,  particularly  the 
liver  and  spleen,  which  become  rich  in  iron.  In  acute  anemia  pro- 
duced by  hemolytic  sera,  with  destruction  of  more  than  half  the  blood 
in  three  days,  nearly  all  the  iron  from  the  destroyed  hemoglobin  can 
be  found  in  the  liver,  spleen  and  kidneys,  there  being  but  little  lost 
through  the  urine  even  in  so  severe  an  anemia  as  this  (INIuir  and 
Dunn).*^"  Excretion  of  bile-pigments  increases,  and  ^^hematogenous 
jaundice"  may  result,  the  bile-pigments  that  are  present  in  the  blood 
being  derived  from  the  hematoidin  of  the  hemoglobin  molecule. 
Changes  in  metabolism  occur  which  are  quite  similar  to  those  ob- 
served in  other  forms  of  anemia,  with  fatty  changes  in  all  the  paren- 
chymatous organs,  increased  protein  katabolism,  and  an  excessive 
quantity  of  pigmentary  substances,  particularly  urobilin,  in  the  urine. 

CHLOROSIS 

The  characteristic  feature  of  the  blood  in  chlorosis  is  the  rela- 
tively small  amount  of  licmoglobin  in  pro])()rtion  to  the  number  of 
corpuscles.     Apparently,  therefore,  the  fault  lies  rather  in  the  manu- 

40  See  Mohr,  Zeit.  exp.  Path..  lOOfi   (2),  4.35. 
40aGrafe,  Deiit.  Arch.  klin.  Med..  11)1,'>   (US),  14S. 

41  Virchow's  Arch.,   1864    (20),  241. 
4iaA8h.  Arch.  Int.  Med.,   1014    (14),  8. 

41b  Drinker  and  Ilurwitz,  Arch.  Int.  Med,  I'M.')  (If)),  73:];  Jour.  Exp.  yied., 
1915    (21),  401. 

4ic  Jour.  Patli.  and  I'-act..  IDlf)   (10),  417. 


cuiJtuosL^  303 

facture  of  hemoglobin  than  in  cither  a  destruction  or  a  deficient 
formation  of  red  corpuscles.  Er])cn's '-  analyses  of  chlorotic  blood 
showed  that  the  total  amount  of  protein  is  decreased,  chiefly  because 
of  the  deficiency  of  hemoglobin;  the  relation  of  serum  globulins  and 
serum  albumins  is  unchanged,  wliile  tlie  proportion  of  fi])rinogen  is 
increased.  There  is  much  more  fatty  substance  than  normal  in  both 
the  serum  and  the  erythrocytes,  but  the  lecithin  is  decreased  both 
in  the  serum  and  in  the  total  blood,  although  somewhat  increased  in 
the  red  cells.  Cholesterol  is  decreased  in  both  serum  and  corpuscles. 
In  the  asli,  pliosphoric  acid,  potassium,  and  iron  are  decreased,  w-hile 
calcium  and  magnesium  are  both  increased.  An  apparent  increase  in 
sodium  chloride  exists,  but  it  is  only  apparent,  being  the  result  of 
the  increase  in  the  proportion  of  plasma  in  the  blood.  The  total 
amount  of  plasma  is  greatly  increased    (polyplasmia). 

The  decrease  in  hemoglobin  is  demonstrable  chemically  as  well  as 
microscopicall}',  Becquerel  and  Rodier  *^  having  found  the  amount  of 
iron  in  the  total  blood  decreased  in  direct  proportion  to  the  apparent 
decrease  in  hemoglobin,  which  frequently  falls  to  30^0  per  cent.,  and 
may  drop  to  20  per  cent,  or  possibly  less.  Alkalinity,  as  determined 
by  titration,  is  diminished  in  some  cases,  but  generally  remains  nearly 
rionnal.  The  corpuscles  are  said  to  contain  a  larger  proportion  of 
water  than  normal,  independent  of  the  proportion  of  water  present 
in  the  serum.  Limbeck  found  their  isotanicity  {i.  e.,  the  strength  of 
NaCl  necessary  to  prevent  hemolysis)  veiy  low — about  0.38-0.4  per 
cent.  NaCl. 

Very  few  changes  seem  to  occur  in  the  organs  of  the  body;  the 
usual  tendency  to  lay  on  fat,  and  the  occurrence  of  fatty  degenera- 
tion observed  commonly  in  anemias,  may  be  exhibited,  and  are  cor- 
related with  Erben's  observation  of  an  increased  fat  content  in  the 
blood;  but  these  changes  are  often  absent.  The  hypoplasia  of  the 
aorta,  upon  which  Virchow  laid  so  much  stress,  is  now  considered  to 
be  of  little  or  no  significance.  Thrombosis  is  a  not  infrequent  com- 
plication of  chlorosis,**  and  is  probably  favored  by  the  increased 
platelet  and  fibrin-content  of  the  blood  and  the  tendency  to  fatty 
changes  in  the  vessel-walls. 

Studies  of  nitrogenous  mctaholism  by  Yannini  *°  showed  practically 
no  alterations  except  a  slight  retention  of  nitrogen. 

Etiology. — As  to  the  etiology  of  chlorosis,  chemical  findings  indi- 
cate some  possibilities  and  negative  others,  but  decide  nothing.  That 
chlorosis  does  not  depend  upon  a  hemolytic  poison  is  well  established 

*2Zeit.  klin.  ^led.,  1002  (47),  302.  See  also  Frohmaier.  Folia  Hematol.,  1015 
(20),  115:  Boumer  and  Burger,  Zeit.  exp.  Path..  1013   (13),  351. 

*3  For  literature  see  Krehl,  "Basis  of  Symptoms,"  lOlfl,  p.  106;  Ewing,  "Clinical 
Pathology-  of  the  Blood,"  1001.  p.  167:  Kossler,  Cent.  f.  inn.  Med.,  1807   (18),  657. 

**  See  Schweitzer.  Vircliow's  Arch.,  1898  (152),  337,  and  Lcichtenstern,  Miinch. 
med.  Woch.,  1809    (46),  1603. 

*5Virchow's  Arch.,  1004   (176),  375. 


304  DISTURBANCES    OF    CIIiCULATIOX 

hy  the  following  facts :  there  is  no  free  hemoglobin  in  the  blood 
plasma,  and  even  less  iron  in  the  serum  ash  than  normal ;  lecithin  and 
cholesterol,  important  products  of  disintegration  of  erythrocytes,  are 
both  decreased  in  the  serum ;  hematogenous  icterus  does  not  occur,  and 
the  amount  of  pigments  in  the  urine  and  feces  is  decreased. 

Apparently,  therefore,  hematogenesis  is  at  fault,  particularly  the 
formation  of  hemoglobin,  since  this  is  more  deficient  than  is  the  total 
number  of  red  corpuscles.  The  rapid  improvement  in  the  condition 
that  follows  the  administration  of  iron  would  seem  to  indicate  that  a 
deficient  supply  of  iron  is  the  cause  of  chlorosis,  but  numerous  ob- 
jections exist  to  this  hypothesis.  Bunge  advanced  the  idea  that  under 
normal  conditions  the  only  form  of  iron  that  can  be  absorbed  is  that 
which  is  combined  with  proteins,  particularly  nucleoproteins ;  iron  ad- 
ministered in  inorganic  form,  or  as  compounds  with  organic  acids, 
he  believed,  can  all  be  recovered  from  the  feces,  and,  therefore,  is  not 
absorbed.  He  suggested  that  in  chlorosis  the  iron  taken  with  the 
ordinary  food  is  precipitated  in  the  intestines  by  sulphides  or  other 
products  of  intestinal  putrefaction,  and  hence  there  results  a  de- 
ficiency in  the  amount  of  iron  absorbed  and  available  for  the  manu- 
facture of  hemoglobin.  The  inorganic  iron  given  in  chlorosis,  Bunge 
believes,  owes  its  efficiency  to  its  saturating  all  of  these  sulphides  so 
that  the  nucleoprotein-iron  is  not  precipitated,  and  can,  therefore,  be 
absorbed.  ]\Iany  objections  have  been  raised  to  Bunge 's  hypothesis, 
however,  for  competent  observers  have  failed  to  find  any  abnormal 
putrefaction  in  chlorosis,  and  others  have  found  that  sulphide  of  iron 
itself  gives  good  results  in  the  treatment  of  chlorosis,  while  bismuth 
and  other  sulphur-binding  substances  are  without  effect.  Further- 
more, Bunge 's  contention  that  iron  administered  in  medicinal  form  is 
not  absorbed  seems  to  have  been  completely  disproved  by  several  ex- 
perimenters.^** 

As  a  consequence  of  all  these  conflicting  data  we  are  at  present 
completely  in  the  dark  as  to  the  reason  for  that  failure  properly  to 
manufacture  hemoglobin  which  seems  to  be  at  the  bottom  of  chlorosis. 
The  hypothesis  that  iron  and  arsenic  favor  recoveiy  by  stimulating 
the  hemogenetic  tissues,  which  is  urged  by  v.  Noorden  and  others,  is 
unsatisfactory  in  the  extreme,  and  explains  nothing.  There  is  abso- 
lutely no  question  tluit  administration  of  iron  restores  the  composi- 
tion of  the  blood  to  normal,  usually  quite  rapidly,  and  this  seems  to 
leave  as  most  probable  the  explanation  that  in  some  way  an  iron 
starvation  is  the  fundamental  cause  of  cldorosis.  However,  as  Ewing 
says,  any  theory  must  be  inadequate  that  fails  to  take  into  account  the 
age  of  puberty,  tlie  female  sex,  and  tlio  function  of  menstruation. 

40  Full  review  witli  biljliofjiapliv  bv  E.  Clever,  Krpebiiisse  Phvsiol..  1005  (.5), 
698;  Meinertz,  Cent,  riiysiol.  u.  Path".  StolTwedi.,  lOOT   (2),  652." 


PERNICIOUS  ANEMIA  305 

PERNICIOUS  ANEMIA 

111  contrast  to  chlorosis  many  evidences  of  hematolysis  may  be 
found  in  pernicious  anemia,  particularly  the  increased  amounts  of 
iron  in  the  liver,  spleen,  and  kidneys;  hemoglobinemia  and  hemoglo- 
binuria; increase  in  ur()l)ilin,  and  not  infreqneiitly  icterus. 

Chemical  Changes. — l-'rlicn's  i"  analyses  of  tlie  blood  in  ])ornici()us  anemia  prave 
the  f(>ilo\\inu-  losiilts:  Tlic  proteins  are  decreased,  both  in  the  sernm  *'a  and  in 
the  blood  as  a  whole:  part ieularly  in  the  latter,  because  of  the  <.'reat  decrease  in 
the  number  of  corpuscles.  The  <|uantity  of  proteins  in  the  individual  corpuscles 
is  increased,  correspondintj  to  their  increased  size.  Fibrin  is  decreased  in  total 
amount,  but  is  relatively  normal  as  compared  with  the  total  proteins:  alliuniin  is 
normal ;  serum  olobulin  much  decreased.  The  proportion  of  water  is  much  in- 
creased, both  in  the  serum  and  in  tlie  corjiuscles.  Fat  is  present  in  normal 
amounts;  cholesterol  is  decreased,  althoujjh  in  relatively  normal  quantities  in 
the  corpuscles.  Lecithin  is  decreased  in  the  total  blood,  but  increased  propor- 
tionately in  the  corpuscles.  The  total  ash  is  increased,  owin<r  chiefly  to  an 
excessively  lar^e  jiroportion  of  XaCl  and  a  slight  increase  in  calcium  and  map- 
jiesiimi ;  potassium  and  ])hosphoric  acid  are  decreased  because  f)f  the  small  num- 
ber of  corpuscles:  but  the  serum  itself  contains  more  P^jO.  and  potassium  than 
normal.  Although  the  total  iron  is,  of  course,  much  decreased,  there  is  iron  in 
the  serum  (indicating  hemolysis)  and  the  proportion  of  iron  in  the  corpuscles  is 
increased;  but  as  the  amount  of  iron  in  the  corpuscles  is  even  greater  than  cor- 
responds to  the  hemoglobin  increase,  it  would  seem  that  either  the  hemoglobin 
in  pernicious  anemia  is  very  rich  in  iron,  or  that  the  corpuscles  c(mtain  iron 
bound  in  some  form  other  than  hemoglobin. 

The  analyses  of  Rumpf  ^^  agree  quite  closely  with  those  of  Erben,  and.  taken 
jointly  with  other  analyses  in  the  literature,  show  the  large  proportion  of  Mater 
in  the  blood,  the  small  amount  of  solids,  the  large  amount  of  NaCl.  and  the 
decrease  in  potassium  and  iron.  Rumpf  also  examined  the  brain,  liver,  heart, 
and  spleen  in  one  ease.  Water  was  found  increased  in  the  heart,  decreased  in 
the  other  organs,  the  solids  not  being  decreased  in  any  of  the  organs.  Tiiere  was 
little  fat  in  any  of  the  organs  or  in  the  blood,  but  NaCl  was  generally  increased. 
The  liver  contained  four  or  five  times  as  much  iron  as  normal :  the  spleen  three 
or  four  times.  Rumpf  is  inclined  to  lay  great  stress  on  the  general  jioverty  of 
the  body  in  potassium,  and  suggests  its  therapeutic  application.  Svllaba  *" 
found  bilirubin  and  also  free  hemoglol)in  in  the  l)lood  of  seven  patients.  Fowell  so 
found  a  considerable  excess  of  iron  in  the  blood  over  tlie  amount  combined  with 
hemoglobin.  Schimim  5i  could  lind  no  proteoses  or  other  evidences  of  protein 
decomposition  in  the  blood  in  a  case  of  pernicious  anemia,  but  he  did  find  free 
hematin.sia  The  tendency  to  hemorrhage  observed  in  this  disease  may  dejiend  on 
a  slight  decrease  in  the  prothrombin  and  a  reduction  in  the  numl)er  of  platelets. ''ib 

V.  Jaksch  and  also  v.  Limbeck  ^>^  have  foiuid  some  decrease  in  total  alkalinity, 
which  probably  depends  on  the  loss  of  proteins  and  their  fixed  alkali. ■'''•''  The  red 
corpuscles  are  very  susceptible  to  hemolysis  by  lowering  of  osmotic  pressure 
("high  isotonicity,"  equal  to  0.;')4  per  cent.  XaCl — v.  Limbeck).  The  specific 
gravity  of  the  whole  blood  is,  of  course,  decreased,  but  the  corpuscles  themselves 

*"  Zeit.  klin.  :Med.,  1900   (40),  266.     Reumer  and  Riirger.  Zeit.  exp.  Path..  1013 
(13),  343. 
*"a  See  also  Heudorfer,  Zeit.  klin.  IMed.,  1!)1.3    (70),  103. 
■t^Rerl.  klin.  ^Yoch..  1001    (38),  477. 
40  Abst.  in  Folia  Ilematnl..   1004    (1),  2S3  and  580. 
•'i'l  Quart.  Jour.  :Med..  1013    (6).  170. 
■"■1  Ilofmeister's  Reitr.,   1003    (4).  4r)3. 
siaZeit.  physiol.  Chem.,  1016   (07),  32. 
■"■ih  Drinker  and  Hurwitz,  Arch.  Int.  Med.,  1915    (15),  733. 

52  "Klin.  Pathol,  des  Blutes,"  Jena.  1896,  p.  311. 

53  See  Brandenburg,  Zeit.  klin.  Med..  1902    (45),  157. 

20 


306  DISTURBANCES    OF    CIRCULATIOX 

have  practically  iioinial  specilic  gravity,  while  tlie  decrease  is  chielly  in  the 
serum. 54 

111  six  cases  of  pernicious  anemia  Stiihlen  ^r.  found  ahundant  iron  in  the  liver 
and  spleen  microscopically,  and  less  constantly  in  the  kidneys  and  bone-marrow. 
Hunter  '"J  gives  the  following  results  of  analysis  of  the  liver,  kidney,  and  spleen 
for  iron: 

Liver  and 
kidney.  Spleen. 

Pernicious  anemia,  seven  cases  average  .  .  .  0.360  per  cent.  0.125  per  cent. 
Other  conditions    (with  anemia),  average     .     ".   0.079         "  0.3(52         " 

Healthy  organs 0.084         "  0.090 

Iron  is  also  fouiul  in  the  hemolymph  glands,  sometimes  more  abundantly  than 
in  the  spleon    (\\'arthin)  .57 

Extensive  studies  on  the  protein  mctaholism  of  pernicious  anemia  by  Rosen- 
quist  58  showed  tliat  there  is  a  considerable  destruction  of  tissue  proteins,  as 
indicated  by  nitrogen  loss,  but  that  at  times  nitrogen  may  be  stored  up  for  brief 
periods.  At  times  there  may  also  be  an  excessive  elimination  of  purine  nitrogen, 
indicating  destruction  of  nuclear  elements.  Calorimetric  studies  show  the  metab- 
olism to  be  slightly  above  normal. 5'<a  in  anemia  due  to  liothriocephaliis  quite 
similar  changes  were  observed. 

Hunter  59  describes  the  condition  of  the  urine  in  pernicious  anemia,  particularly 
with  reference  to  the  elimination  of  much  "pathological  urobilin,"  "o  which  seems 
to  be  produced  by  intracellular  destruction  of  hemoglobin.  Iron  may  also  appear 
in  the  urine  in  increased  quantities. ^i 

Summary. — Putting  together  the  above  findings,  we  see  that  in 
pernicious  anemia  we  have  every  evidence  that  excessive  hemolysis 
is  taking  place,  and  the  fact  that  continued  poisoning  by  toln.ylendia- 
mine  *^-  and  other  hemolytic  poisons,  such  as  that  of  Bofhrlocephalus, 
may  give  rise  to  a  condition  resembling  pernicious  anemia  ver^'  closely, 
indicates  strongly  that  hemolytic  poisons  are  the  case  of  pernicious 
anemia.  Histological  studies  show  the  same  thing,  and,  as  Warthin  ^"^ 
says:  "The  hemolysis  of  pernicious  anemia  does  not  differ  in  kind 
from  that  occurring  normally  or  in  certain  diseased  conditions;  the 
difference  is  one  of  degree  only."  The  hemolysis  seems  to  go  on 
chiefly  inside  of  phagocytic  cells  instead  of  in  tlie  blood,  probably 
because  the  phagocytes  pick  up  the  corpuscles  as  soon  as  they  have 
been  injured  by  the  hemolytic  poisons.  In  some  instances  choles- 
terol administration  improves  the  anemia,  which  suggests  that  the 
poison  attacks  the  lipoids  of  the  corpuscles,"*  as  so  many  hemolytic 

54  Bonninger,  Zcit.  exp.  Path.,  1912   (11),  1. 

55  Deut.  Arch.  klin.  Med.,  1S95    (54),  248   (literature). 

58  Lancet,  1903  (i),  283;  similar  re.-;ults  obtained  bv  Uvfl'i-l.  .Tour.  Path,  and 
Bact.,  1910   (14),  411. 

57Amer.  Jour.  Med.  Sci.,  1902   (124),  074. 

58Zeit.  klin.  Me<l.,  1903  (49),  193  (literature).  See  also  :Min()t.  Bull.  .Johns 
Hopkins  Tlosp.,  1914    (25),  338. 

581.  Mcver  and  DnP.ois,  Arcli.  ln(.  :\Ied.,  191()  (17).  90.'):  (.'rate.  Deut.  Arcli. 
klin.  :Med.,  1915   ( 1 18) ,  148. 

5!)  British  Meil.  Jour.,  1890    (ii),   1   and  81. 

«f' See  also  Mott,  Lancet,  189(»  (i),  287;  and  Svllal)a,  .\bst.  in  Volia  lleiuatol.. 
1904    (1),  283. 

"1  Kennerkneclit,  Virchow'a  Arch.,  1911  f205).  89.  Not  conlirmed  bv  Q\ieckeii- 
stedt,  Zeit.  klin.  :\red.,  1913    (79),  49;  bildiography. 

«2Svllaba,  Ihniter    (lor.  cit.) . 

•'4Sw  Keicher,  Berl.  klin.  Woch.,  1908   (45),  1838. 


LEUKEMIA  307 

a^'eiits  do.  Ihthriotcphalus  anemia,  which  so  closely  resembles  the 
"pernicious''  form,  seems  to  be  caused  by  a  hemolytic  lipoid,®^  pre- 
sumably a  ehoh'sterol  ester  of  oleic  acid,  and  there  is  a  g^rowing  tend- 
ency to  associate  hemolytic  lipins  witli  the  etiology  of  pernicious 
anemia."''  However,  although  in  liemol^'tic  anemias  tliere  is  an  in- 
creased amount  of  unsaturated  lipins  in  the  blood,  Medak  "^  did  not 
find  the  isolated  lipoids  to  be  particularly  hemolytic.  (See  Hemolysis, 
Chapter  viii.)  The  origin  and  the  nature  of  the  specific  hypothetical 
poisons  have  been  sought  in  vain.  Some  authors  have  referred  them 
to  infections  of  unknown  nature,  occurring  perhaps  in  the  mouth  and 
gastrointestinal  tract  (Hunter),''"  or  to  hemolytic  products  of  intes- 
tinal putrefaction,"-  or  to  faulty  metabolism.  Many  others,  with  per- 
haps the  best  of  grounds,  would  ascribe  pernicious  anemia  to  a  multi- 
plicity of  causes,  which  produce  a  protracted  slight  hemolysis  that 
continues  until  the  hematogenetic  organs  give  out,  their  exhaustion 
being  perhaps  hastened  by  the  influence  of  the  toxic  substances  in  the 
blood;  hematogenesis  then  becomes  insufficient  to  replace  the  lost 
corpuscles,  and  the  picture  of  pernicious  anemia  is  established.'^'* 

LEUKEMIA 

In  leukemia  the  chemical  changes  in  the  red  corpuscles  take  a  less 
prominent  position,  resembling  either  those  of  a  secondary  anemia 
or  chlorosis,  while  the  enormous  number  of  leucocytes  is  the  prominent 
feature  and  causes  marked  alterations  in  the  composition  of  the  blood. 
Large  quantities  of  nucleoproteins  and  also  of  the  intracellular 
en/ymes  are  introduced  into  the  blood  by  the  excessive  leucocytes.  As 
the  leucocytes  are  constantly  breaking  down,  more  or  less  of  the 
products  of  their  decomposition  are  present  in  the  blood  and  appear 
in  the  urine.  Because  of  the  relatively  slight  metabolic  activity  of 
the  Ijonphocytes  the  various  chemical  alterations  are  all  less  marked 
in  IjTnphatic  than  in  myelogenous  leukemia.''"     There  is  a  notable  re- 

65  Tallquist,  Zeit.  klin.  Med.,  1907  (61),  427:  Arch.  exp.  Path.  u.  Pharm..  100? 
(57).  367. 

66  See  Liidke  and  Fejes,  Deut.  Areh.  klin.  :Med.,  1913    (109),  433. 
63  Biochem.  Zeit.,  1914   (59),  419. 

67  Lancet,  1903   (1),  283. 

68  See  Kiilhs  (Arch.  exp.  Path.  u.  Pharm.,  1906  (55),  73),  who  found  tlie  in- 
testinal contents  of  patients  with  chronic  intestinal  disorders  to  contain  lienio- 
lytic  substances  of  undetermined  character.  Hemolytic  lipoids  in  tlie  intestinal 
contents  have  been  described  by  Berber  and  Tsuchiga  (Deut.  Arch.  klin.  Med.. 
1909  (96),  252)  and  Liidke  and' Fejes,  loc.  cit.;  but  this  observation  failed  of  con- 
firmation by  Ewahl    (Deut.  med.  Woch.,  1913    (39),  1293). 

Herter  (Jour.  Biol.  Chem.,  1906  (2),  1)  suggested  a  relation  between  intestinal 
infection  with  B.  aerogenes  capsulatus,  which  produces  hemolytic  substances,  and 
pernicious  anemia. 

69  See  also  Bunting.  .Johns  Hopkins  Hosp.  Bull.,  1905  (16),  222;  Pappenheim. 
Folia  Serologica.  1910   (10),  217. 

"0  Stern  and  Eppenstein  have  observed  that  the  striking  proteolytic  power  of 
the  leucocytes  from  the  blood  in  myelogenous  leukemia  is  not  shown  by  the 
leucocvtes  in  Ivmphatic  leukemia  (Sitz.  d.  Schles.  Ges.  f.  vaterliind.  Kultur, 
June  29,   1906)". 


308  DISTURBANCES    OF    CIRCULATION 

duction  ill  antibody  production  in  leukemia,""''  presumablj^  because  of 
the  changes  in  the  bone  marrow ;  it  is  said  that  typhoid  infection  in 
leukemics  may  fail  to  result  in  agglutinin  formation. 

Chemistry  of  the  Blood. — Considering  the  quantitative  alterations  in  the  con- 
stitUL-nts  of  tlie  lihxxl,  wo  find  the  specific  gravity  lowered,  Ijut  not  so  much  as 
it  would  be  in  a  simple  anemia  with  equally  low  hemoglobin,  for  tlie  loss  of 
hemoglobin  is  partly  compensated  by  the  increase  in  leucocytes  and  their  products. 
Fibrinogen  is  usually  increased  in  myelogenous  leukemia.'i  Tlie  serum  sliows 
but  slight  change  in  specific  gravity,  a  slight  decrease  in  proteins  "la  lieing  com- 
pensated by  an  increase  in  the  NaCl.  Tlie  freezing-point  of  the  lilood  is  lowered 
(Cohn'-),  which  is  probably  due  to  the  increase  in  crystalloidal  products  of 
cellular  decompositlun.  Erben  "3  found  that  in  lymphatic  leukemia  the  serum 
contains  less  cholesterol  tlian  normal,  although  the  fat  content  may  be  rather 
high.  Calcium  is  frequently  found  increased,  probably  because  of  destruction 
of  the  bone  tissue.  In  the  red  corpuscles  the  proportion  of  iron,  protein  and 
potassium  is  decreased  as  is  also  that  of  the  cholesterol,  that  of  the  lecithin 
and  water  being  somewhat  increased.  The  total  amount  of  potassium  and  iron 
in  the  blood  is  decreased,  but  the  PjO-,  in  the  ash  is  increased  because  of  tiie  large 
amount  of  nucleoprotein  in  the  blood.  A  number  of  the  earlier  writers  describe 
a  decreased  alkalescence  which  probably  is  due  to  the  deficiency  in  the  fixed 
alkali  of  the  proteins.  There  is  an  increased  excretion  of  iron  in  the  urine  and 
feces.'* 

The  poor  coagulation  of  leukemic  blood  has  long  been  known,  but 
the  reason  for  it  has  not  yet  been  ascertained.  Some  investigators 
have  reported  a  deficiency  in  fibrin,  while  others  have  found  it  in- 
creased. More  recent  reports,  however,  indicate  that  there  is  no 
marked  change  in  either  the  amount  of  fibrinogen  or  of  the  fibrin- 
ferments.  Erben  '^^  found  a  normal  amount  of  fibrin  in  the  blood  in 
lymphatic  leukemia;  and  in  three  cases  of  myelogenous  and  one  of 
lymphatic  leukemia,  Pfeiffer  "  found  the  amount  of  fibrinogen  nearly 
normal.  This  is  quite  remarkable  in  view  of  the  fact  that  in  ordinary 
forms  of  leucocytosis  both  the  amount  of  fibrinogen  and  the  rapidity 
of  clotting  are  increased.  It  is,  therefore,  extremely  difficult  to  under- 
stand the  poor  coagulability  of  leukemic  blood,  but  study  of  the  fac- 
tors of  coagulation  by  modern  methods  may  clear  this  up,  for  in  one 
ease  so  studied  Whipple  '^^  found  an  increase  in  antithrombin. 

Decomposition  Products. — Of  particular  interest  is  the  finding 
in  the  blood  of  decomposition  products  of  the  leucocytes,  which  are 
probably  produced  by  autolysis  of  the  leucocytes.  (See  Leucocytic 
Enzymes,  Chapter  iii.)  Normal  leucocytes  are  rich  in  autolytic 
enzymes,  which  under  ordinary  circumstances  seem  to  be  held  in 
check  by  the  antienzymes  of  the  blood.     In  leukemia  this  antienzyme 

TOaKotkv,  Zent.  inn.  Med.,  1014   (35),  9r),3. 

-1  Erben^,  Zeit.  klin.  Med.,  1008  (6G),  278;  full  details  on  composition  of  the 
Idood   in   leukemia. 

71a  Little  change  was  found  in  the  protein  content  of  the  serum  b\  UciKlorfcr, 
Zeit.  klin.  Med.,  1913   (70),  103. 

T2:Mitteil,  aus  dem  Crenzgeb.  ]\led.  u.  Chir.,  1000   (15),  11.  1. 

73  Zeit.  klin.  Med.,  1000   (40),  282. 

74  Kennerknecht,  Vircliow's  Arch..   1011    (205).  80. 
7.-.  Cent.  f.  inn.  Med.,  1904    (25),  800. 

76  Arch.  Int.  Med.,  1913   (12),  637. 


LEUKEMIA  309 

action  seems  to  be  insufficient  to  prevent  leucocytic  autolysis,  for 
even  in  freshly  drawn  l)lood  proteoses  (or  at  least  nou-coa<iulable  pro- 
teins) may  be  present.'"  Aceordin<r  to  Erben,  tliis  is  tr-ue  only  of 
rayelog-enous  leukemia,  the  fresh  blood  in  lymphatic  leukemia  not  only 
being:  free  from  non-eoag:ulable  protein,  but  furthermore  this  product 
of  proteolysis  does  not  soon  develop  when  the  blood  is  kept  aseptically 
at  incubator  temperature.  This  is,  of  course,  what  one  would  expect 
in  view  of  the  well-known  enzyme-richness  of  the  polymorphonuclear 
leucocytes  and  the  scarcity  of  proteolytic  enzymes  in  lymphocytes. 
Erben  states  that  the  neutrophile  cells  seem  to  be  the  chief  source  of 
proteoses,  since  their  granules  soon  disappear  in  blood  that  is  undergo- 
ing: autolysis,  whereas  the  eosinophiles  preserve  their  granules  well,  and 
true  proteoses  are  not  present  in  blood  rich  in  mast  cells  (i.  e.,  mye- 
loma) .  The  marrow,  spleen  and  lymph  grlands  are  found  strongly  pro- 
teolytic (according:  to  the  plate  method),  in  myelogenous  leukemia,  but 
in  lymphatic  leukemia  and  pseudoleukemia,  only  the  marrow  shows  a 
slight  activity.'^  Schnmm  ''^  found  in  the  blood  in  a  case  of  myelogen- 
ous leukemia  several  varieties  of  proteoses,  most  abundant  being  the 
so-called  deutero-albumose ;  in  another  he  also  found  peptone,  leucine, 
and  tyrosine.  In  addition  he  demonstrated  the  autolytic  nature  of 
the  changes  that  occur  in  leukemic  blood  after  death  (see  also  "Auto- 
lysis in  Leukemia,"  Chap.  iii).  Most  observers  have  failed  to  find 
alhumose  in  the  urine  in  leukemia.  Because  of  the  involvement  of 
the  bone  marrow,  small  amounts  of  Bence-Jones  protein,  as  well  as 
Morner's  body,  may  be  found  in  the  urine.®"  Kolisch  and  Burian  ^^ 
not  only  found  nucleoprotein  constantly,  and  albumose  frequently, 
but  in  one  case  of  lymphatic  leukemia  they  found  histon  in  the  urine, 
which  undoubtedly  came  from  nucleoprotein  decomposition. 

The  oxidase  reaction  is  conspicuous  in  certain  of  the  cells  of  mye- 
loid leukemia,  especially  the  large,  non-granular  cells  of  acute  leu- 
kemia,®^ but  it  is  not  known  that  these  oxidases  influence  the  chem- 
istry of  the  disease.  In  spite  of  the  richness  of  leucocytes  in  lipases 
the  serum  shows  no  increased  lipolytic  activity. ^^-"^ 

Protein  Metabolism. — Stejskal  and  Erben  ®^  studied  the  metabolism 
of  a  case  of  myelogenous  and  of  a  case  of  lymphatic  leukemia,  and 
found  the  nitrogen  loss  much  greater  in  the  myelogenous  form,  al- 
though food-absorption  was  better  than  in  the  lymphatic ;  they  con- 
sider that  protein-destroying  forces  are  at  work  in  myelogenous 
leukemia,  similar  to  those  of  cancer  cachexia,  so  that  nitrogenous  equi- 
librium cannot  be  attained. 

7TFor  literature  see  Erben,  Zeit.  f.  Keilk.    (Int.  Med.  Abt.),  in03    (24),  70. 

•8  Jochmann  and  Zioo^ler,  Miineh.  med.  Woch.,  1906    (.53),  200.1. 

-n  Hofmeister's  Beitr.,  1003    (4),  442:   Dout.  med.  Woeh.,  lOOo    (31),  183. 

soBofrfrs  and  Oiithrie.  Bull.  .Tolins  Hopkins  llosp.,  1013   (24),  308. 

S2  Zeit.  klin.  Med.,  1806    (29).  374    (literature  on  albuminuria  in  leukemia). 

Ri  Dimn,  Quart.  Jour.  Med.,  1013   (6).  203. 

siararo.  Zeit.  klin.  Med.,  1913    (78),  286. 

83  Zeit.  f.  klin.  Med.,  1900   (39),  151. 


310  DISTURBANCES    OF    CIRCULATION 

As  the  most  characteristic  products  of  decomposition  of  nucleo- 
proteins  are  the  purine  bases,  one  would  also  expect  to  find  them 
present  in  leukemia,  and  early  writers  mention  the  finding  of 
purine  bases  and  uric  acid  in  the  blood  and  spleen.  The  urinary 
finding's  in  this  respect  have  been  very  variable.  Ebstein  ®*  observed 
the  complication  of  leukemia  with  gout  which  he  considered  a  coin- 
cidence, and  also  noted  uric-acid  concretions  in  the  urinary  passages 
in  four  cases.  Numerous  other  authors  have  described  increased  uric- 
acid  elimination,  while  some  have  observed  increase  in  the  purine 
bases,  either  witli  or  without  uric-acid  increase.  ]Magnus-Levy  ®^ 
observed  a  particularly  large  uric-acid  output  in  acute  leukemias,  but 
also  found  that  the  relation  between  the  number  of  leucocytes  and 
the  uric  acid  is  extremely  variable.  Sometimes  the  nitrogen  loss  is 
very  great — even  as  much  as  20  gm.  per  day — and,  corresponding 
with  the  destruction  of  nucleoproteins  and  the  resulting  uric-acid 
formation,  phosphoric-acid  excretion  is  often  greatly  increased — even 
up  to  15  gm.  per  day.  On  the  other  hand,  the  results  obtained  by 
many  other  writers  have  been  in  every  respect  extremely  variable ; 
some  have  found  no  increase  in  uric  acid,  some  even  report  a  decrease ; 
likewise  the  PoO-,  has  been  found  even  less  than  normal.  For  ex- 
ample, in  a  carefully  studied  case  of  lymphatic  leukemia,  Henderson 
and  Edwards  ^*^  found  during  six  months  no  excessive  excretion  of 
uric  acid  or  phosphoric  acid.  Zalesky  and  Erben  found  likewise  no 
considerable  increase  in  the  uric  acid  in  lymphatic  leukemia,  but  in 
myelogenous  leukemia  the  uric  acid  was  much  increased ;  on  the  other 
hand,  the  amount  of  elimination  of  purine  bases  was  reversed  in  the 
two  forms,  and  creatin  M^as  decreased  in  both.  Lipstein  ^'  found  no 
excessive  elimination  of  amino-acids  even  in  myelogenous  leukemia. 
An  increase  in  calcium  is  quite  constantly  observed,  and  attributed  to 
the  bone  destruction  ^^  occurring  in  this  disease. 

Undoubtedly  these  variations  in  results  depend  upon  the  known 
fluctuations  in  the  course  of  the  pathological  processes  of  leukemia  ;  the 
number  of  leucocytes,  the  size  of  the  lymphatic  organs,  and  the  general 
condition  of  the  patient  all  vary  greatly  from  time  to  time,  often  with 
remarkable  rapidity,  and  the  excretion  of  products  of  metabolic  ac- 
tivity must  vary  likewise.  It  can  hardly  be  questioned  that  the 
enormous  increase  in  the  amount  of  lymphoid  tissue  in  the  body  and 
blood  must  give  rise  to  a  greatly  increased  nuclein  catabolism,  with 
conse(iuent  appearance  of  its  products  (uric  acid,  purine  bases,  and 
phosphoric  acid)  in  the  urine.  This  seems  to  be  well  demonstrated 
by  the  increased  elimination  of  uric  acid  and  purine  bases,  together 
with  a  general  increase  in  the  nitrogen  output  that  has  been  frequently 

84  For  litoraturo  soo  rr'smiH'  bv  Walz  in  Cent.  f.  I'atliol..  lOOl    (12).  985. 

85  Virchow's  Arch.,   ISOS    (152).   107. 
soAmer.  Jour,  of  Physiol.,   1!)03    (0),  417. 
87  Loc.  cit.  inf. 


LEUKEMIA  311 

observed  following  the  therapeutic  use  of  a:-rays  in  leukemia,  which 
is  attributed  to  the  increased  autolysis  that  x-rays  are  known  to  pro- 
(hu'e.'''*  According  to  Kosensterii  "*''  the  x-rays  affect  chiefl.v  the  leuco- 
genic  tissues  rather  than  the  adult  leucocytes.  Lipstein  '■"'  also  found 
an  excessive  elimination  of  amino-acids  in  the  urine  of  leukemic  pa- 
tients treated  by  x-rays.**^  According  to  Curschmann  and  Gaupp,^- 
the  bk)od  of  leukemic  patients  who  liave  been  exposed  to  x-rays  con- 
tains a  specific  leucocytotoxin,  whicli  may  be  produced  by  a  process  of 
autoimmunization  against  the  leucocytic  substance  set  free  by  the  dis- 
integrated leucocytes.  Capps  and  Smith  ^'^  have  obtained  similar  re- 
sults. 

Charcot's  crystals  (also  called  Charcot-Les'den  and  Charcot-Xeumann  crystals) 
represent  a  peculiar  and  strikino;  product  of  nuclear  destruction  that  has  fre- 
quently been  found  associated  with  leukemia.  These  crystals  were  first  observed 
by  Robin ''4  (1853)  in  leukemic  tissues,  but  have  been  named  after  Charcot,  who, 
with  Kobin,  described  tlieir  properties.  They  were  described  by  Charcot  as  color- 
less, retractile,  elongated  octahedra;  insoluble  in  alcohol,  ether,  and  glycerol; 
soluble  in  hot  water,  acids,  and  alkalies;  size  variable,  from  0.016  by  0.005  mm.  up. 
These  crystals  have  been  found  not  only  in  the  tissues  and  blood  of  cadavers,  but 
also  occasionally  in  the  freshly  drawn  blood  of  leukemics.  Poehl  03  believes  them 
to  be  the  same  as  Buttcher's  spermin  crystals,  and  derived  from  decomposed 
nucleins.  Schreiner  considers  that  these  spermin  crystals  are  phosphoric  acid 
salts  of  spermin    (C2H5X),  or,  as  Majert  and  Schmidt  give  it,  C^HioN,,  with  the 

structure  HX  <;pTT- pTr"  >  XH,  thus  being  similar  to,  although  not  identical 

with,  piperazin.  The  entire  question  of  the  composition  of  spermin  is  still  im- 
settled,9o  however;  and  it  is  probable,  furthermore,  that  the  crystals  found  in 
leukemia  are  not  identical  with  the  crystals  observed  in  semen. 

Crystals  that  appear  similar  are  also  found  in  asthmatic  sputum,  empyema, 
and  ascites  fluid,  bone-marrow,  and  tumors,  and  it  has  been  suggested  that  they 
are  derived  from  or  related  to  the  oxyphile  granules  of  the  eosinophiles.9"  This 
view  implies  an  agi'eement  with  Gumprecht's  opinion  that  the  crystals  seen  in 
bone-marrow,  asthmatic  sputum,  etc.,  are  not  spermin,  but  of  protein  nature. 
As  can  be  seen,  the  nature  and  significance  of  Charcot's  crystals  are,  at  the  pres- 
ent time,  quite  undetermined. 

Summary. — The    chemical    changes   observed   in   leukemia   depend 

ss  Radiiim  also  causes  marked  metabolic  changes  in  leukemia,  with  enormously 
increased  excretion  of  urea,  ammonia  and  total  X.  and  especially  of  P^0_ ;  never- 
theless tlie  increase  in  uric  acid  excretion  is  slight  (Ordwav,  Knudson  and  Erdos, 
Boston  :Nred.  and  Surg.  .Jour..  1917   (176),  490). 

89Miinch.  med.  Woch.,  1906    (53),  1063. 

90  Hofmeister's  Beitr..  1905    (7),  527. 

91  Literature  on   effects   of  x-ravs  in  leukemia,   see  Arneth.   Berl.   klin.   Wocli., 

1905  (42),  1204:  Musser  and  Eds'all,  I'niv.  of  Penn.  Med.  Bull..  1905  '18),  174: 
Rosenberger.    Miinch.    med.    Woch..    1906     (53).    209:    Williams,    Biocheia.    Jour.. 

1906  (1).  249;  Lossen  and  IMorawitz,  Deut.  Arch.  klin.  Med.,  1905  (S3),  288; 
Koniger,  Deut.  Arch.  klin.  Med.,  1906    (87).  31. 

92Munch.  med.  Woch.,  1905    (52),  2409. 

93  Jour.  Exp.  Med.,  1907  (9),  51;  see  also  Klieneberger  u.  Zoeppritz.  ^liinch. 
med.  Woch.,  190G  (53),  Xo.  18:  Milchner  u.  Wolfi".  Berl.  klin.  Woch.,  1906  (43). 
No.  23. 

94  Literature  given  Ijv  v.  Levden,  Festschrift  fiir  Salkowski.  Berlin,   1904,  p.   1. 
95Deiit.  med.  Woch.. "1895    (21).  475. 

90  Literature,  see  Hammarsten,  Amer.  Transl.,  1904,  p.  420. 

97  Literature,  see  Floderer,  Wien.  klin.  Woch.,  1903  (16),  276;  Predtetschcnskv, 
Zeit.  klin.  Med.,  1906    (59),  29. 


312  DISTURBANCES    OF    CIRCULATION 

upon  the  excessive  quantity  of  leucocytes  and  lymplioid  tissue,  which 
undergo  processes  of  disintegration  at  irregular  intervals,  with  the 
result  that  the  products  of  nucleoprotein  destruction  (uric  acid, 
purine  bases,  and  phosphoric  acid)  appear  in  the  urine  in  increased 
quantities.  As  the  large  neutrophiles  contain  abundant  autolytic 
enzymes,  the  products  of  cell  autolysis  (proteoses,  amino-acids,  and 
products  of  nucleoprotein  destruction)  may  appear  at  times  in  the 
urine  and  in  the  blood;  hecause  of  the  small  amount  of  such  enzymes 
in  the  lymphocytes,  these  changes  are  all  much  less  marked  in  lymph- 
atic leukemia.  Charcot's  crystals,  which  are  perhaps  derived  from 
leucocytic  nucleoproteins,  may  be  found  in  the  blood  and  tissues. 
The  changes  in  the  red  cells  are  chiefly  those  of  a  secondary  anemia, 
with  occasionally  some  chlorotic  features.  The  chemical  findings  of 
leukemia  throw  no  light  whatever  upon  the  cause  of  the  disease. 

Pseudoleukemia  and  Hodgkin's  disease  show  only  the  evidences  of  a 
secondary  anemia,  without  the  chemical  changes  of  either  leukemia  or 
pernicious  anemia.  There  seems  to  have  been  little  study  of  the 
chemical  processes  of  these  diseases.  Moraczewski  ^*  reports  a  study 
of  metabolism  in  one  case,  designated  by  him  as  pseudoleukemia  and 
so  quoted  in  subsequent  literature,  although  the  only  leucocyte  count 
mentioned  in  the  original  article  was  171,000.  This  case  showed  some 
retention  of  nitrogen  and  calcium,  with  little  change  in  the  phosphorus 
and  purine  bases  in  the  urine. 

HYPEREMIA 

ACTIVE  HYPEREMIA 

This  condition  is  associated  with  but  few  chemical  changes.  Cer- 
tain chemicals  may  cause  active  hyperemia ;  some  locally,  as  in  the 
case  of  irritants,  such  as  alcohol,  ether,  ammonia,  mustard,  etc.,  which 
act  either  by  producing  a  local  vasodilator  stimulus  or  by  paralyzing 
the  vasoconstrictors.  Other  substances  may  produce  active  hyperemia 
in  special  vascular  areas,  e.  g.,  cantharides  causes  active  hyperemia  in 
the  kidneys,  probably  because  of  its  elimination  through  these  organs ; 
pilocarpin  causes  active  hyperemia  in  the  salivarj^  glands  and  skin, 
which  is  associated  with  increased  function.  In  general,  functional 
activit.y  is  associated  with  active  hyperemia,  and  Gaskell  ^  has  suggested 
that  this  is  due  to  atonicity  of  the  vascular  muscle,  the  result  of  de- 
creased alkalinity  of  the  lymph  flowing  away  from  the  active  organ 
along  the  vessel-walls,  it  having  been  found  that  alkalies  cause  a  tonic 
contraction  and  acids  an  atonic  dilation  of  arterial  muscle.^'' 

osVirdiow's  Arch.,  ISOS   (151),  22. 

1  Quoted  by  Lazanis-Barlow,  "]\Tamial  of  General  Patliolo<iry."  1904,  p.  120. 
lii  Sec  discussion  l)v  \\'oolIeY,  Jovir.   Anier.  Med.  Aaaoc,   1914    (63),  2279;   and 
bv  Adler,  Jour.  Pharm.,  1910  "(8),  297. 


PASSIVi:  NYI'EKEMIA  313 

Pathological  active  hyperemia  is  seldom  of  long  enough  duration 
to  lead  to  any  alterations  in  the  tissues  in  which  it  oceurs.  The  blood 
itself  remains  unehanged,  except  that  the  venous  blood  going  from  the 
part  contains  nuich  less  CO,  and  more  oxygen  tliaii  usual,  because 
more  oxj-gen  is  brought  to  the  tissues  than  can  be  used.- 

PASSIVE  HYPEREMIA 

Passive  hyperemia  is  almost  equally  unassociated  with  chemical 
changes,  especially  in  its  etiology,  which  depends  almost  solely  upon 
mechanical  factors.  Some  chemical  alterations  result,  however,  from 
the  changes  in  the  stagnating  blood,  which  may,  if  the  obstruction 
to  outflow  is  severe,  become  of  venous  character  in  the  capillaries  of 
the  congested  area.  Oxidation  in  the  tissues  is,  therefore,  impaired, 
and  some  fatty  changes  may  result,  e.  g.,  in  the  center  of  congested 
liver  lobules.  Waste  products  accumulate,  and  possibly  noxious 
products  of  metabolism  are  formed  under  lack  of  oxidation ;  either 
from  these  causes  or  solely  from  pressure  and  lack  of  nutrition  there 
is  a  tendency  to  atrophy  of  the  more  specialized  parenchymatous  cells, 
and  a  proliferation  of  connective  tissues.  The  atrophy  of  parenchyma 
is  seen  particularly  in  the  liver,  the  increase  of  connective  tissue  in  the 
spleen.^  In  the  kidney  neither  atrophy  nor  stroma  proliferations 
are  pronounced,  but  the  renal  function  is  greatly  impaired,  since  it 
depends  upon  the  amount  and  quality  of  the  blood  brought  to  the 
kidney.*  Whether  connective-tissue  proliferation  in  hyperemia  de- 
pends upon  overnutrition  or  upon  irritation  by  waste-products,  or  is 
compensatory  to  parenchymatous  atrophy,  may  be  looked  upon  as 
still  an  open  question.  Probably  only  the  first  two  factors  apply  to 
the  connective-tissue  growth  observed  in  the  congested  spleen,  the 
clubbing  of  the  fingers  in  congenital  heart  disease,  or  the  thickening 
of  the  subcutaneous  tissues  in  passive  congestion  of  the  lower  ex- 
tremities. 

Changes  in  the  Blood. — Venous  blood  differs  from  arterial,  not 
only  in  its  increased  load  of  COo  and  other  waste  products,  but  also 
in  other  ways.  Venous  blood  generally  clots  less  readily  than  arterial 
blood.^  It  contains  more  diffusible  alkali  because  the  CO,  combines 
with  and  tears  away  part  of  the  bases  that  are  held  by  the  proteins, 
especially  in  the  corpuscles,  and  so  alkaline  carbonates  are  formed 

2  Polycythemia  {Vaquez-Osler  disease)  is  accompanied  by  an  increase  in  tlie 
total  nitrocren  of  the  blood,  in  proportion  to  the  number  of  erythrocytes;  but  the 
nitrogen  content  of  the  individual  erythrocyte  is  decreased,  (v.  .Jaksch,  Zent.  inn. 
Med.,  1912   (33),  397). 

3  See  Christian,  Jour.  Amer.  Med.  Assoc,  1905    (45),  1615. 

4  See  Ro^yntree  and  Geraghty,  Arch.  Int.  Med.,  1913  (11),  121;  Xonnenljruch, 
Deut.  Arch.  klin.  Med.,  191,3   (110),  162. 

sVierordt  (Arch.  f.  Heilk.,  1878  (19),  193)  found  coagulation  faster  in  the 
blood  in  passive  congestion  than  in  normal  venous  blood;  but  Hascbrock  (Zeit.  f. 
Biol.,  1882  (18),  41)  found  that  if  the  stasis  is  protracted,  the  coagulation  be- 
comes delayed  because  of  the  excess  of  CO;. 


314  DISTURBANCES    OF    CIRCULATION 

and  enter  the  plasma.  Blood  from  the  jng'ular  vein  on  this  account 
contains  20-25  per  cent,  more  diffusible  alkali  than  carotid  blood 
(Hamburger).®  Since  the  bactericidal  power  of  the  blood  has  been 
shown  to  increase  directly  with  the  alkalinity,  this  property  may  be 
of  importance  in  pathology.  For  example,  the  relative  infrequency 
of  infections  in  the  right  side  of  the  heart  may  not  depend  solely  upon 
lessened  liability  to  endocardial  damage,  as  generally  considered,  but 
is  possibly  due  in  part  to  the  greater  bactericidal  power  of  venous 
blood.  The  same  property  probably  explains  the  favorable  results 
obtained  in  the  treatment  of  local  infections  by  artificially  produced 
passive  congestion."  Too  severe  a  stasis,  with  resultant  edema,  prob- 
ably favors  local  infection.'''^ 

V.  Fodor  ^  found  that  animals  surviving  infections  show  an  in- 
creased blood  alkalinity,  w^hereas  in  those  that  died,  the  alkalinity 
was  decreased ;  also,  he  found  the  resistance  increased  by  intravenous 
injections  of  alkalies.  Other  observers  °  have  noted  a  decrease  in  re- 
sistance after  injecting  acids  into  the  blood.  According  to  Calabrese, 
the  alkalinity  of  the  blood  increases  in  immunization  of  animals 
against  toxins,  while  Cantani  found  the  injection  of  toxin  followed 
by  a  decrease  in  alkalinity.  Hamburger  has  shown  that  the  bac- 
tericidal power  of  the  blood  may  be  increased  in  vitro  by  shaking  it 
with  CO2  as  a  result  of  the  increased  alkalinity,  aided,  perhaps,  by 
some  slight  bactericidal  power  of  the  CO,  itself;  he  also  found  the 
blood  more  strongly  bactericidal  in  venous  congestion  than  normally, 
and  the  lymph  from  a  congested  part  was  also  found  more  strongly 
bactericidal  than  normal  lymph.  Hamburger  ^^  has  also  found,  how- 
ever, that  eheraotaxis  is,  if  anything,  slightly  decreased  under  the  in- 
fluence of  COo,  as  also  is  phagocytosis:  large  amounts  of  CO,  may 
reduce  the  phagocytic  power  for  coal  particles  by  25-50  per  cent. 
Hamburger's  results  as  to  the  bactericidal  power  of  human  blood  in 
venous  stasis  have  been  confirmed  by  Laqueur."  Schiller  ascribes  this 
not  to  increased  alkalinity,  but  to  disintegration  of  leucocytes  with 
liberation  of  bactericidal  substances.^^*^ 

The  blood  in  the  veins  and  capillaries  in  passive  congestion  is  gen- 
erally richer  in  corpuscles  than  normal,  ])erhaps  because  of  some  loss 
of  water,^-  although  this  is  not  constant,  applying  particularly  to 
more  recent  or  more  local  processes;  in  long-continued  stasis,  as  in 
congenital  heart  disease,  the  blood  may  be  diluted. ^^     In  the  eoncen- 

<■>  Vircliow's  Arch.,  1899   (156),  329;  also,  "OsmotisHuT  Dnick  \iiid  loiu'iilehre." 

7  Roo  Bier,  "Hvpersemie  als  Heilmittel,"  Leipsic,  190;]. 
7ii  Glasewald,  Cent.  Grenz.  Med.  Cliir.,  1915    (18),  50". 

8  Cent.  f.  15akt.,  1890    (7),  753. 

0  Literature,  see  Hanilmrfjer   (loc.  citfi),  p.  281. 

10  Vircliow's  Arch.,  1899    (156).  329. 

11  Zoit.  exp.  Patli.  u.  Therap.,  1905    (1),  670. 
uaBeitr.  klin.  Cliir..  1913   (84),  IT.  1. 

i-^CIrawitz,  Dent.  Arch.  f.  klin.  Med.,  1895    (54),  588. 
13  See  Krehl,  "Pathologische  Pliysiologie,"  1904,  p.  201. 


Tiii{(>  ]fiiosis  315 

trated  blood  of  passive  congestion  the  corpuscles  may  number  six  to 
eight  millions  per  .cubic  millimeter,  while  the  concentration  of  the 
solids  of  the  serum  may  be  at  tlie  same  time  reduced  CKrehl).  The 
viscosity  of  such  blood  is  higher  than  that  of  normal  l)lood.^*  In 
acute  stasis  the  proi)orti()n  of  serum  proteins,  especially  the  albumin, 
increases  with  the  duration  of  the  stasis ;  no  changes  occur  in  the  non- 
protein constituents  of  the  blood  (Rowe).^^^ 

THROMBOSIS 

The  cliemistry  of  thrombosis  in  most  respects  resolves  itself  into  the 
chemistry  of  fibrin  formation,  a  subject  which  is  so  extensively  con- 
sidered in  most  treatises  on  physiological  chemistry  and  physiology 
that  it  does  not  seem  desirable  to  give  here  anything  more  than  the 
essential  principles  involved  in  the  clotting  of  the  blood,  as  now  under- 
stood, as  an  introduction  to  the  consideration  of  the  same  process  as 
it  occurs  under  pathological  conditions.  In  spite  of  innumerable  in- 
vestigations, our  knowledge  of  the  actual  participants  and  processes 
involved  in  the  formation  of  fibrin  is  in  a  veiy  unsatisfactory  and 
fragmentary  state.  Some  facts  seem  well  established,  however,  and 
we  have  a  general  idea  of  the  subject  that  may  be  applied  with  ad- 
vantage to  the  consideration  of  thrombosis. 

FIBRIN   FORMATION.i- 

Several  difFeront  substances  seem  to  l)e  eoneerned  in  the  forniation  of  fibrin, 
of  -which  tlie  first  of  importance  is  its  antecedent,  fibrinogen.  Fibrinogen  is  a 
simple  protein,  related  to  the  globulins,  and  diflfering  chiefly  in  its  icady  coagula- 
bility, not  only  by  fibrin  ferment,  but  also  by  heat,  salts,  and  other  coagulating 
agencies.  By  itself,  however,  it  shows  no  tendency  to  coasrulate  spontaneously. 
According  to  Goodpasture.ifi  fibrinosen  is  formed  through  tlie  combined  activity 
of  the  liver  and  intestines,  although  earlier  writers  have,  variously  described  its 
formation  in  the  bone  marrow,  leucocytes,  liver  or  intestines.  The  amount  of 
fibrinogen  present  in  the  blood  is  actually  quite  small,  the  fibrin  formed  in  nor- 
mal clotting  being  but  0.1  to  0.4  per  cent,  of  tlie  weiffht  of  the  blood.  Acted 
upon  by  the  fibrin-ferment,  it  yields  the  characteristic  insoluble  protein  fibrin,  in 
crystalline  form  under  favorable  conditions,!"  but  we  do  not  know  definitely  what 
changes  the  fibrinogen  undercroes  in  this  process.  Fibrin  resembles  in  its  insolu- 
bility the  proteins  coagulated  by  heat,  alcohol,  etc.,  but  when  kept  aseptically  for 
some  time,  it  becomes  asain  dissolved;  this  process  of  fibrinoh/sis  probably  de- 
pends upon  proteolytic  enzymes,  which  fibrin,  in  common  with  other  sulistances 
of  similar  physical  nature,  has  the  property  of  dragging  out  of  solution  and 
holding  firmly.  Undonljtedly  entangled  leucocytes  are  also  an  important  factor 
in  the  fibrinolysis. lo  which  is  greatly  increased  in  phosphorus  poisoning  and  when 
the  liver  is  excluded  from  the  circulation,  a  fact  suggesting  that  tlie  liver  may 
form  inhibiting  substances. 

i4Determann.  Zeit.  klin.  :Med..  inOO    (.5n),  U.  2-4. 

14a  .Tour.  Lab.  flin.  :\fed.,  1016   (1),  48.5. 

15  For  literature  and  full  discussion  see  Hammarsten's  or  ^lathews'  Physiolog- 
ical Chemistry;  ;Morawitz,  Ergebnisse  der  Phvsiol.,  Abt.  1,  1004  (4),  307.  and 
the  andbuch  "d.  Biochem.,  1008  II  (2),  40; 'Leo  Loeb,  Biochem,  Centr..  1907 
(6),  829. 

ifi  Amer.  Jour.  Physiol.,  1914   (3.3),  70. 

1'  See  Howell.  Amer.  Jour.  Phvsiol.,  1014  (3.5),  143;  Hekma,  Internat.  Zeit. 
physik.  chem.  Biol.,  191.5    (2),  279. 

19  See  Morawitz,  loc.  cit.;  also  Pvulot,  Arch,  internat.  d.  Physiol..  1904  (1),  152. 


316  DISTURBAyCEl^    OF    CIRCULATION 

Theories  of  Fibrin  Formation. — The  great  problem  is  the  nature  and  the  place 
and  manner  of  uriuin  of  the  librin-forming  enzyme,  generally  called  fihrin-ferment 
(also  ])lasmasc.  thrombin  and  coa<iuUn).  Tlie  most  fimdamental  theory  of  the 
origin  and  nature  of  fil)rin-fernient  is  that  of  Alexander  Schmidt,  which  may  be 
briefly  described  as  follows:  The  ferment,  Schmidt  believes,  exists  in  the  plasma 
in  an  inactive  (prozyme  or  zymogen)  form,  which  he  calls  prothrombin.  Upon 
disintegration  of  the  leucocytes  there  is  set  free  a  substance,  which,  acting  upon 
the  prothrombin,  converts  it  into  the  active  thrombin;  this  activating  agent 
Schmidt  designates  as  the  zymoplasiic  substance.  With  various  modifications 
this  stands  to  the  present  day  as  a  basic  theory. 

It  having  been  sliown  that  calcium  facilitates  the  formation  of  fibrin,  Pekel- 
haring  advanced  tlie  idea  that  the  protliruinl)iii  does  not  exist  in  the  plasma,  but 
is  liberated  from  the  leucocytes,  and,  coml)ining  with  the  calcium  of  the  plasma, 
forms  the  thrombin.  Morawitz  considers  three  substances  necessary  for  tlie  forma- 
tion of  thromliin.  (1)  the  prothrombin  or  thrombogen,  which  he  believes  orig- 
inates in  the  blood-plates;  (2)  the  zymoplastic  substance  or  thrombokinase,  which 
is  liberated  from  the  leucocytes  into  the  plasma:  (3)  calcium  salts.  Howell, 20 
however,  explains  coagulation  as  follows:  Circulating  blood  normally  contains 
all  the  necessary  factors  for  fibrin  formation,  i.  e.,  fibrinogen,  prothrombin  and 
calcium.  But  there  is  also  present  an  inhiliiting  substance,  antithrombin,  which 
j)revents  the  calcium  from  activating  the  protliromljin  into  thromliin.  In  shed 
blood  there  appears  a  thromboplastin,  derived  from  the  platelets  or  the  tissues, 
wliich  neutralizes  the  antithrombin  and  thus  permits  thrombin  to  form.  Rett- 
ger  22  holds  that  the  coagulation  of  the  blood  is  not  a  true  enzyme  action  at  all, 
while  Bordet  and  Delange  23  consider  that  thrombin  is  formed  by  the  interaction 
of  cytozyme  from  the  platelets  or  tissue  cells,  and  serozyme  of  the  plasma. 
Mathews  follows  Wooldridge  and  considers  the  clotting  of  the  blood  as  essentially 
the  crystallization  of  a  phospholipin-protein  compound,  blood  plasma,  the  stability 
of  which  compound  is  easily  upset  in  many  ways.  The  fibrin  threads  are  essen- 
tially liquid  crystals  coming  out  of  a  saturated  solution,  tlie  blood  plasma,  which 
is  practically  a  dilute  protoplasm.  It  will  not  serve  our  purpose,  however,  to  go 
further  into  the  hypotheses  and  disputes  over  these  questions,  which  are  detailed 
more  fully  in  the  literature  previovisly  cited,  but  it  may  be  stated  tJiat  numero\is 
American  observers  have  found  Howell's  theory  to  fit  well  Avith  both  experimen- 
tal and  clinical  observations  on  the  variations  in  the  coagulability  of  the  blood. 

The  question  has  been  raised  as  to  whether  the  leucocytes  or  platelets  secrete 
their  fibrin-forming  constituent  (be  it  thrombokinase  or  prothrombin  is  a  matter 
of  minor  importance  to  the  pathologist)  or  liberate  it  only  after  their  disin- 
tegration. So  far  as  jiathological  processes  go,  the  latter  seems  to  be  the  case, 
the  disintegration  apparently  occurring  whenever  the  leucocytes  come  in  contact 
with  a  foreign  body  or  witli  dead  and  injiu'ed  tissues.  Tlie  stroma  of  red  cor- 
puscles also  contains  tlirombokinase.^i  Of  the  substances  that  may  be  isolated 
from  tissues,  eephalin  is  found  especially  active  in  producing  thrombosis,  and 
may  be  related  to  or  identical  with  the  thrombo]ilastin.2-'J 

Tissue  Coag'ulins. — Among  the  other  points  that  are  of  importance  in  patho- 
logical conditions  is  the  fact  that  not  only  the  leucocytes,  but  also  tissue-cells, 
can  liberate  fibrin-forming  substances  (coaguUns  is  the  non-committal  term  ap- 
plied by  Loeb).  Howell  considers  that  the  eflfect  of  the  tissue  "coa'j;ulins"  is 
merely  to  neutralize  tlie  antithromliin  of  the  blood,  if  such  coagulins  actually 
exist;  possilily  tliere  is  tliromlio])histin  in  the  tissues.  These  coagulating  agents 
are  present  in  tissue  extracts  and  are  liberated  whenever  the  tissues  are  injured; 
muscle  is  rich  in  coagulin,  as  are  also  tlie  liver  and  kidney,  and.  wliich  is  par- 
ticularly important,  the  blood-vessel  wall  (L.  Loeb).  Pieces  of  these  tissues 
placed  in  contact  with  fibrinogen  solution  will  bring  about  prompt  clotting.  An- 
other important  fact  is  that  the  coagulins,  whetlier  derived  from  leucocytes  or 
from  the  tissues,  have  a  cci-tain  degree  of  specificity — that  is,  they  act   solely  or 

20Amer.  .Tour.  Phvsiol..   1911    (20),   187. 
22Amer.  Jour.  Pliysiol.,  100!)   (24),  40(i. 

23  Ann.  Inst.  Pasteur..  1912  (20),  (ioT.  See  also  Lee  and  \'iiicent.  Arch.  Int. 
Med.,  1914    (1:5).  3!).S. 

24  Barratt,  .Tour.  Path,  and  Bact.,  191.3   (17),  30:{. 

2.'-.  Howell,  Aiiicr,   .[(.ur.    I'livsiol.,    1912    ( .'{l  ) ,    1,  MacLean,   ibid..    191(1    (41),  2."')0. 


COAdULATIOX   or  THE  BLO(Jj  317 

most  rapidly  with  fil)rinofrcn  of  Ijlood  of  tlio  species  from  wliicli  tiiey  are  ob- 
tiiiiu'd.'-''  Jii  some  iiistanci's  this  spccilicity  is  ahsoliito,  hut  inoie  jieiieraliy  (  par- 
ticuhirly  in  tlio  maiiuiuilia  )  it  is  only  relative.  J.oeh  also  found  tliat  tiie  amount 
of  tissue  ooaj>ulin  was  not  decreased  in  or<;ans  altered  by  phospliorus  jioisoning, 
althou^di  duriufj  experimental  autolysis  the  coajrulins  disajjpear.  Wiien  tissue 
coagulins  and  blood  coaiiulins  act  together,  the  elTect  is  greater  than  the  sum 
of  their  independent  actions,  indicating  the  probability  that  they  combine  in 
some  way  to  [jroduce  a  particularly  active  coagulin.  The  blood  eoagulins  are 
(piite  dili'erent  from  the  tissue  eoagulins  in  many  important  respects,  and  the 
eoagulins  cannot    be  l(M)ked  ujion  as  a  single  substance  of  dillVrcnt   origins. 

Blood-platelets. —  It  is  still  undetennined  just  what  jiart  the  platelets  ])lay  in 
coagulation.  The  well-known  observation  that  in  thrombosis  the  fibrin  is  often 
first  formed  about  masses  of  platelets  clinging  to  the  wall  of  the  ve.ssel  indicates 
that  they  participate  in  the  process,  and  Bizzozero  and  others  have  maintained 
that  the  platelets  and  not  the  leucocytes  are  the  source  of  the  prothrombin. 
Numerous  studies  on  the  relation  of  the  platelets  to  disease  conditions  have  in- 
dicated a  certain  parallelism  l)et\veen  their  number  and  the  tendency  to  coagulation 
observed  in  the  various  diseases  (Welch).  Howell  I)elieves  the  platelets  to  be 
the  chief  source  of  thromboplastin,  which  neutralizes  the  antithrombin  of  the 
blood  and  tims  causes  clotting.  Eordet  and  Delange  consider  the  ])latelets  of 
more  importance  than  the  leucocytes  in  producing  participant*  of  the  coagulat- 
ing mechanism.  The  histological  evidence  of  the  importance  of  the  ])latelets  in 
thrombus  formation  is  conclusive  (see  Zurhelle,  Derewenko),  and  Cramer  and 
Pringle  -«  state  that  coagulation  cannot  occur  without  platelets.  Kemp  29  con- 
cludes, from  a  thorough  review  of  the  subject,  that  the  blood-platelets  are  visually 
normal  or  subnormal  in  number  during  acute  infectious  diseases,  but  increase 
rapidly  if  the  disease  terminates  by  crisis:  in  pernicious  anemia  the  number  is 
always  greatly  diminished,  although  in  secondary  anemias  they  may  sometimes 
be  increased;  in  purpura  lurmorhagica  the  number  of  plates  is  enormously  di- 
minished, which  is  perhaps  related  to  the  slowness  of  the  clotting  of  the  i)lood 
in  this  condition.  Duke  3'^  states  that  when  the  i)latelet  count  falls  below  10,(100 
per  cubic  mm.  there  is  delayed  coagulation  and  a  tendency  to  purpura :  with 
counts  above  40,000  tliere  is  usually  no  hemorrhagic  tendency.  If  the  platelet 
count  is  reduced  artificially  (by  benzene,  diphtheria  toxin)  a  similar  purpuric 
tendency  is  observed.  Poisons  that  in  large  doses  reduce  the  platelet  count,  will 
increase  it  if  in  small  doses. 

Calcium  Salts. — The  exact  significance  of  calcium  in  fibrin  formation  is  still 
imsettled.  Blood  from  which  the  calcium  has  been  precipitated  will  not  coagu- 
late, and  the  addition  of  calcium  salts  v'\\\  promptly  cause  it  to  do  so.  The  vari- 
ous hypotheses  advanced  to  explain  the  way  in  which  calcium  influences  the 
clotting  process  are  not  in  agreement.  One  hypothesis  is  tliat  the  calcium  ions 
are  necessary  for  the  transformation  of  prothrombin  into  thrombin  (Pekelharing, 
Hammarsten,  IMorawitz),  the  thrombin  consisting  of  a  compoimd  of  prothrombin, 
calcium  salts,  and  thrombokinase.  Howell  considers  that  no  kinase  is  necessary, 
the  calcium  activating  the  prothrombin  whenever  it  is  not  inhibited  by  anti- 
thrombin. 

Modification  of  Coagulability. — Another  important  matter  for 
consideration  is  the  etit'eet  of  various  substanees  in  moclifyin<>-  the  rate 
or  completeness  of  the  coagulation  of  the  blood.  In  the  first  place,  we 
have  the  well-known  fact  that  if  blood  is  drawn  into  a  glass  vessel 
well  coated  with  oil  or  vaseline,  through  a  cannula  similarly  ]n-otected, 
no  coagulation  will  take  place ;  but  if  any  unoiled  foreign  stibstance 
enters,  even  particles  of  dust,  coagulation  begins  at  once.     The  ex- 

20  Leo  Loeb,  Univ.  of  Penn.  Med.  Bull..  1004   (16),  .3S2 ;  Muraschew.  Deut     \rch 
klin.  Med.,  1904   (SO).  187. 

28  Quart.  Jour.  Exper.  Physiol.,  1913    (6),  1 

29  Jour.  Amer.  ^Med.  Assoc.,  in06    (46).   1022. 

30  Jour.   Exp.  Med.,    1011     (14).   265;    Arch.   Int.   :\red.,    1012    (10).    44.1:    Jour 
Amer.  Med.  Assoc,  1915   (65),  1600. 


318  DISTVRBAyCES    OF    CIRCULATION 

planation  is  that  the  leucocytes  do  not  liberate  their  coagulating  sub- 
stances until  they  have  been  injured  by  contact  with  some  foreign 
body,  and  the  experiment  proves  the  importance  of  this  action  of  the 
leucocytes,  as  well  as  explaining  wliy  the  blood  does  not  coagulate  dur- 
ing life.  The  classical  experiment  of  the  ligation  of  a  vein  without 
injury'  to  the  endothelium,  which  permits  the  blood  to  remain  stag- 
nant for  a  long  period  without  clotting,  depends  upon  the  same  fact, 
namely,  that  normal  endothelium  neither  liberates  coagulin  itself  nor 
injures  the  leucocytes  so  that  they  disintegrate.  Loeb  recalls  the 
observation  of  Overton  that  lipoids  are  important  constituents  of  the 
cell  membranes,  and  suggests  a  similarity  between  the  vessel  lining 
and  the  oiled  cannula,  but  analyses  of  aortic  endothelium  have  shown 
a  rather  low  lipin  content  (8.41-9.25  per  cent.),  although  peritoneal 
endothelium  has  much  more  (13  to  15  per  cent.).^^  The  suggestion 
that  the  vessel  walls  contain  an  anti-coagulin  could  not  be  confirmed  by 
Loeb.  Since  leucocytes  are  constantly  undergoing  disintegration  in 
the  blood  and  tissues  under  normal  conditions,  it  might  be  asked  why 
they  do  not  cause  clotting  then  and  there.  In  explanation  Loeb  ad- 
vances his  observation  that  the  coagulins  are  destroyed  during  cell 
autolysis,  and  suggests  that  when  leucocytes  normally  disintegrate,  the 
coagulins  are  first  destroyed  by  autolysis.  It  has  also  been  shown  that 
the  cells  and  serum  contain  substances  which  inhibit  or  prevent  coagu- 
lation, and  it  is  possible  that  these  play  an  important  part  under  nor- 
mal conditions  in  preventing  coagulation  by  products  of  cell  disintegra- 
tion, much  as  other  antienzymes  are  supposed  to  act  in  preventing 
autodigestion  of  living  cells. 

Coagulation  of  drawn  blood  may  be  retarded  experimentally  by  re- 
moval of  the  calcium  by  precipitation  as  oxalate,  fluoride,  etc. ;  also  by 
diminishing  the  oxygen  and  increasing  the  COo,  by  addition  of  solu- 
tions of  neutral  salts  in  large  amounts,  l)y  diluting  greatly  with  water, 
or  by  keeping  the  blood  cold.  Coagulation  may  be  hastened  by  moder- 
ate heat,  by  whipping,  exposure  to  air,  by  contact  with  much  foreign 
matter,  and  by  the  addition  of  watery  extracts  from  many  different 
tissues  and  organs.  Poisons  that  destroy  the  ])latplets  reduce  the 
coagulation  (Duke).  Of  particular  interest  i)ath<)l()gically  is  the  re- 
tardation of  coagulation  that  follows  injections  of  proteoses  (the  so- 
called  "peptone"  solutions)  and  also  various  other  protein-containing 
solutions,  such  as  organ  extracts,  bacterial  toxins,  snake  venoms,  eel 
serum,  extract  of  leeches  or  of  Vneinarm,  impure  nucleo-protein  solu- 
tions, or  solutions  of  various  colloids.  ^lost  of  tlies(^  substances  (>.  g., 
peptone,  eel  serum)  cause  reduction  of  coagulability  when  injected 
into  animals,  and  are  without  effect  on  blood  removed  from  the  body. 
A  few,  however,  prevent  coagulation  of  di-awn  blood  (snake  venom, 
extract  of  leeches).  When  substances  of  the  first  class  are  injected 
in  sufficient  f(uantities,  there  occurs  first  a  period  of  accelerated  co- 
st Tait.  Quart.  .Iniir.  Exj..  Physiol.,  ini.T    (8),  301. 


COAGULATIOS   OF  THE  ULOOD  319 

agulation  whieli  may,  particularly  in  the  ease  of  org'au  extracts, 
cause  prompt  death  from  intravascular  clotting;  if  the  animal  sur- 
vives, there  follows  a  period  of  decrease  or  total  iiihil)ition  of  co- 
agulability of  the  blood,  both  within  the  vessels  and  after  removal 
from  the  body.  The  first  period  of  increased  coagulability  undoubt- 
edly depends  upon  the  formation  of  a  large  amount  of  fibrin-ferment, 
but  it  has  not  yet  been  satisfactorily  explained  how  the  inhibition  of 
coagulation  is  i)r()duced.  Apparently  the  fibrin-ferment  formed  at 
first  is  rai)idly  destroyed,  but  it  is  thought  by  some  that  it  is  con- 
verted into  a  substance  that  neutralizes  the  fibrin-ferment  that  may 
be  formed  later,  or  that  a  true  anticoagulin  is  formed.  It  is  also 
among  the  possibilities  that  all  the  available  prothrombin  or  throm- 
bokinase  is  used  up  during  the  first  stage  of  acceleration.  As  before 
]nentioned,  the  blood  and  tissues  contain  substances  that  inhibit 
coagulation,  and  it  may  be  that  these  are  secreted  in  excessive 
amounts,  a  view  which  is  receiving  much  support  from  recent  observa- 
tions. According  to  Davis  "'-  injection  of  tlirombin  is  followed  quickly 
by  an  increase  in  the  amount  of  antithrombin  in  the  blood.  It  has 
been  found  that  in  animals  deprived  of  the  liver  no  coagulation- 
inhibiting  substances  are  formed  in  the  blood  after  injection  of  pro- 
teoses, hence  Delezenne  believes  that  the  substances  of  this  class  act 
by  causing  a  destruction  of  leucoc3'tes,  thus  liberating  a  substance 
which  increases  coagulation  and  also  another  substance  retarding  co- 
agulation ;  the  first  of  these  is  destroyed  by  the  liver,  leaving  the  re- 
tarding substance  to  act  unopposed."'  Leech  extract  {hirudin)  pre- 
vents clotting  by  means  of  an  antiferment  action,  combining  with  the 
thrombin.''*  Snake  venom,  however,  acts  upon  the  thrombokinase 
(Morawitz). 

Coagulability  of  the  Blood  in  Disease. — In  disease  the  alterations 
in  the  coagulability  of  the  blood  depend  upon  much  the  same  factors. 
In  all  conditions  associated  with  suppuration  and  leucocytosis  the 
amount  of  fibrinogen  is  increased.  This  is  especially  true  of  pneu- 
monia.^^ The  fluidity  of  the  blood  in  septicemia  is  probably  dependent 
upon  the  appearance  of  the  coagulation-inhibiting  pha.se  that  follows 
the  action  of  the  products  of  cell  destruction,  including  among  them 
proteoses.  In  this  connection  should  be  mentioned  the  observation 
of  Conradi,^*^  who  found  that  among  the  products  of  autolysis  is  a 
coagulation-inhibiting  substance  which  is  not  destroyed  by  heat,  dif- 
fuses readily,  and  in  general  behaves  unlike  the  proteins.     This  or 

32Amer.  Jour.  Physiol.,  1011    (29),  160. 

33  The  manner  in  whieh  gelatin  injections  affect  tlie  blood  coaffulability  is  not 
yet  understood  (see  Boggs,  Deut.  Arcli.  klin.  :\Ied.,  1004  (70),  530)  ;  Moll  (Wien. 
klin.  Woch.,  1003    (16),  1215)    found  an  increase  in  fibrinogen. 

3*  Hirudin  mav  contain  antikinase  (Mellanby,  Jour,  of  Phvsiol..  1000  (38), 
441). 

35Dochez,  Jour.  Exp.  Med.,  1012    (16),  603. 

36  Hofmeister's  Beitr.,  1901    (1),  137. 


320  DISTURBANCES    OF    CIRCULATION 

similar  substances  may  well  play  a  part  in  affecting  coagulation  in 
infectious  diseases,  and  Whipple  ^'  has  found  a  decreased  coagula- 
bility in  sei)ticemia  because  of  the  })resence  of  an  excess  of  anti- 
thrombin.  It  may  also  be  mentioned  that  animals  soon  acquire  an 
immunity  against  proteoses,  so  that  their  inhibiting  influence  is  no 
longer  shown.  This  corresponds  to  the  observation  of  Kanthack  ^^ 
that  immune  serum  against  venom,  neutralizes  very  effectively  the 
anticoagulating  principle  of  venom ;  an  amount  of  antiserum  alto- 
gether insufficient  to  neutralize  the  toxic  properties  of  venom  will 
neutralize  its  property  of  preventing  clotting.  The  bacterial  prod- 
ucts may  also  modify  coagulation,  and  L.  Loeb '"  has  found  that 
different  organisms  are  unequally  effective  in  this  respect,  Staphylo- 
coccus aureus  being  much  more  powerful  in  causing  coagulation  than 
any  others  tested ;  *°  typhoid,  diphtheria,  tubercle,  and  xerosis  bacilli 
and  streptococci  being  without  any  apparent  effect,  while  pyocyan- 
eus,  prodigiosus.  and  colon  bacilli  occupy  an  intermediate  position. 
Furthermore,  after  the  organisms  are  killed  by  boiling,  this  effect  is 
greatly  reduced,  showing  that  it  does  not  depend  merely  upon  the 
mechanical  action  of  the  bacteria,  but  probably  upon  bacterial  prod- 
ucts contained  in  the  culture-media. 

After  phosphorus-poisoning  the  blood  may  become  non-coagulable, 
which  Jacoby  *^  ascribed  to  an  absence  of  fibrinogen  in  the  blood,  be- 
cause of  a  fibrinogen-destroying  ferment  in  the  liver.  Doyon  *-  has 
made  a  similar  finding  in  chloroform  necrosis  of  the  liver,  but  he  at- 
tributes especial  importance  to  an  excess  of  antitlirombin  liberated 
from  the  liver  in  these  conditions.  Whipple  has  also  found  a  de- 
crease in  fibrinogen  with  chloroform  necrosis  and  cirrhosis  of  the 
liver.*-''  In  other  instances  of  decreased  coagulability  the  fibrinogen 
is  present,  generally  in  normal  amounts.  After  death  the  blood  be- 
comes incoagulable  because  the  fibrinogen  is  destroyed  through  a 
process  similar  to  that  of  fibrinolysis;  *^  this  fibrinolysis  may  be  com- 
plete as  early  as  ten  hours  after  death.  The  other  proteins  of  the 
blood  do  not  seem  to  be  corresi)on(lingly  attacked.  Thrombokinase 
is  also  scanty  in  cadaver  blood,  but  there  seem  to  be  no  coagulation- 
inhibiting  substances  present.  In  anaphylactic  shock  the  coagula- 
bility is  reduced  or  abolished,  associated  wath  which  is  a  leucopenia.** 

37  Arch.  Int.  Med..  1012   (9),  305. 
-«  Cited  by  La/.anis-Barlow,  p.  141. 
30  .Tour.  Med.  Hesearcli,  \'M):\   (10),  407. 

41  ]\Tueli  (Biocliem.  Zeit.,  1!»0S  (14),  14:5)  states  that  stai>hyloeoeeus  oontains 
tlirf)nit)()kinase. 

•»i  Zeit.  phvsiol.  Chem.,  1000  (30),  IT;")-,  also  Doyoii  rt  a].,  Compt.  Bend.  Soc. 
Biol.,  1005    (58),  403. 

42  Compt.  Bend.  Soc.  Biol.,  1005  (58),  704;  .lour,  piiys.  et  path.,  101-2  (14), 
229. 

42a  P.ull.  .lohna  Hopkins  Ilosp.,  1013   (24).  207. 

43  Morawitz,  Hofmeister's  Beitr.,   lOOf,    (8).  1. 

44  The  ineoagulaliility  of  menstrual  blood  is  ascribed  to  a  lack  of  lilirin  ferment 


COAGULATION  OF  THE  BLOOf)  321 

Whipple  ^ '  states  that  the  antithrombin-prothrombin  balance  in  the 
■  blood  is  in  delicate  equilibrium,  but  preserved  by  strong  factors  of 
^safety.  The  prothrombin  factor  is  rarely  involved,  most  notably  in 
melena  neonatorum  and  aplastic  anemia,  and  such  conditions  may  be 
relieved  by  injecting  normal  blood,  through  the  added  prothrombin. 
The  antithroinhiu  factor  is  often  excessive  in  hemorrhagic  conditions, 
especially  with  hepatic  injury,  or  it  may  be  lowered  and  lead  to  throm- 
bosis from  relatively  slight  injuries.  Obviously  the  injection  of  nor- 
mal blood  will  harm  rather  than  help  patients  with  hemorrhage  due 
to  excessive  antithrnmbin.  Antithrombin  is  often  found  increased  in 
diseases  of  the  blood-forming  organs,  e.  g.,  leukemia,  possibly  as  a 
reaction  to  the  products  of  disintegration  of  corpuscles;  and  hence 
hemorrhagic  tendencies  are  noted  in  these  diseases.  In  icterus  the 
notable  tendency  to  hemorrhage  seems  to  depend  upon  the  binding  of 
the  calcium  of  the  blood  by  the  bile  pigments,**'  and  administration 
of  calcium  may  bring  the  coagulation  time  back  to  normal  with  a  cor- 
responding decrease  in  the  hemorrhagic  tendency. 

PfeifPer  ^'  estimated  the  fibrin  content  of  the  blood  in  disease,  and 
found  it  increased  in  diseases  with  leucocytosis  (pneumonia,  rheuma- 
tism, erysipelas,  scarlet  fever),  except  leukemia,  where  it  was  normal; 
in  diseases  without  leucocytosis  (typhoid,  malaria,  nephritis),  the 
fibrin  was  normal  in  amount.  Stassano  and  Billon  *^  have,  further- 
more, shown  that  the  amount  of  fibrin-ferment  varies  directly  with 
the  number  of  leucocytes  in  the  blood.  Kollmann  *'-^  found  an  increase 
in  the  fibrin  of  eclampsia,  wdiich  Lewinski  ^°  could  not  substantiate. 
In  experimental  infections  of  animals  Langstein  and  ^Mayer  ^^  found 
a  specific  increase  in  pneumococcus  sepsis,  which  undoubtedly  bears 
an  important  relation  both  to  the  characteristic  fibrinous  nature  of 
the  alveolar  exudate  in  pneumonia,  and  the  striking  amount  of  fibrin 
found  in  pneumococcus  pleuritis,  peritonitis,  etc.  ^Mathews  ^'-  found 
an  increase  in  the  fibrin  with  all  experimental  suppurations. 

The  coagulation  time  of  the  drawn  blood  has  been  the  subject  of 
much  study  by  various  methods,^^  but  as  yet  very  little  agreement  has 
been  obtained.  By  different  methods,  in  which  different  conditions 
for  coagulation  are  presented,  the  normal  coagulation  time  varies  from 
2  to  30  minutes ;  with  most  methods  it  is  5  to  8  minutes.     In  general, 

by  Bell   (Jour.  Path,  and  Bact.,  1914   (18),  462)   and  to  an  excess  of  antitlirombin 
bv  Dienst  (Miinch.  med.  Woch.,  1912   (51),  2709). 

45  Arch.  Int.  Med..  1913   (12),  037. 

46  Lee  and  Vincent,  Arch.  Int.  Med.,  1915  (16),  59. 

.47Zeit.  klin.  Med..  1897    (33),  214:  Cent.  f.  inn.  Med.,  1S9S    (19).  1. 
48Compt.  Rend;  Soc.  Riol.,  1903    (55),  511. 
49  Cent.  f.  Gvniik.,  1897   (21).  .341. 
sopfliitrer's  Arch.,  1903    (100),  611. 

51  Hofmeister's  Beitr..  1903   (5),  69. 

52  Amer.  Jour.  Physiol..  1899   (3),  53. 

53  Full  review  and  bibliography  by  Cohen,  Arch.  Int.  Med.,  1911  (8),  684  and 
820. 

21 


322  DISTUh'BAXCES    OF    CIRCULATION 

coagulability  is  not  constantly  if  at  all  alteral  by  fever,  cancer,  dia- 
betes, slight  secondary  anemias,  or  many  other  diseases,  and  in  nor- 
mal conditions  it  remains  fairly  constant.  In  infants  the  coagulation 
time  is  slightly  shorter  than  in  adults.  The  coagulation  is  hastened 
after  considei-able  hemorrhages,  in  endocarditis,  and  perhaps  in  aneu- 
rism and  thrombosis ;  and  is  commonl}-  delaj'ed  in  the  acute  exan- 
themata, in  hemophilia,  in  purpura  neonatorum,  and  occasionally  in 
some  other  diseases.'*  There  is  entire  lack  of  agreement  concerning 
the  reputed  acceleration  of  coag'ulation  by  oral  administration  of  cal- 
cium salts,  and  retardation  by  citrates;  and  the  supposed  thrombo- 
plastic  influence  of  gelatin  cannot  be  shown  consistently  by  direct  ob- 
servations. In  jaundice,  calcium  salts  probably  have  an  effect,  since 
here  the  cause  of  the  deficient  coagulation  seems  to  be  the  fixation  or 
precipitation  of  the  blood  calcium  by  the  bile  pigments.  It  seems 
probable  that  the  measurement  of  the  time  required  for  coagulation  to 
take  place  in  vitro  does  not  exactly  represent  the  tendency  of  the  same 
blood  to  coagulate  in  the  body  of  the  person  from  whom  it  is  obtained ; 
for  example,  the  injection  of  foreign  serum  has  a  notable  effect  in  stop- 
ping hemorrhages,  but  the  coagulation  time  of  the  recipient's  blood  is 
not  correspondingly  altered.  Whipple's  observations  that  with  a  low 
fibrinogen  content  the  blood  may  coagulate  in  normal  time,  and  yet  the 
clots  be  too  delicate  to  stop  hemorrhage,  explains  at  least  part  of  the 
discrepancy;  and  of  similar  significance  is  the  fact  that  with  a  very 
low  platelet  count  the  blood  may  coagulate  a.s  rapidly  as  normal,  but 
the  clots  do  not  shrink  and  become  firm  (Duke).  Hence  with  a  se- 
vere purpura  hemorrhagica  we  may  have  a  normal  clotting  time. 
In  other  conditions  with  normal  coagulability,  hemorrhages  may  re- 
sult from  excessive  fibrinolysis  which  causes  solution  of  the  clot,  espe- 
cially in  hepatic  diseases.'^'*^ 

THE  FORMATION  OF  THROMBI 

If  we  apply  the  facts  brought  out  in  the  precetling  discussion  rela- 
tive to  the  factors  in  the  coagulation  of  blood,  to  the  manner  and 
conditions  under  which  thrombi  are  formed  in  the  circulating  blood, 
we  find  explanations  for  many  of  the  features  of  thrombosis.  Welch  '^ 
describes  the  steps  in  the  formation  of  a  thrombus  after  injury  to  the 
vessel-wall,  as  follows:  First,  there  is  an  accumulation  of  blood- 
platelets  adhering  to  the  wall   at  the  point  of   injury.     Leucocytes, 

54  See  Dochez,  (.Jour.  Exp.  MckI.,  1912  (16),  69:?),  wlio  found  s.mic  delay  in 
coagulation  in  pneumonia.  Corroborated  by  Minot  and  l.cc,  .lour.  Aiiur.  ^led. 
Assof..  1917   (6S),  545. 

5411  See  (ioodpasture.  Bull.  .Johns  Hopkins  Hosj).,  1914    (25).  :?30. 

•>^>  Albutt's  System,  vol.  (!,  complete  (liscusaion  of  the  {general  featuri's  of  throm- 
bosis; also  see  Kiister,  P^rpeb.  inn.  IMed.,  1913  (12),  667;  Zurhelle,  Ziejrler's 
Beitriifie,  1910  (47),  5.39;  Sdiwalbe,  Krfjebnisse  Pathol. .  1907  (XI  (2)  ).  901; 
I..ubar8ch,  All<r.  Pathol.,  Vol.  1,  Wiesbaden.  1905.  Also  see  Asehofl'.  Ziegler's 
Beitr.,  1912  (.52),  205.  and  Arch.  Int.  Med.,  191;?  (12),  50.1,  eoneerninji  the 
mechanics  of  thrombus  formation. 


FORMATION  OF  THROMBI  323 

wliich  may  l)i'  present  in  small  numbers  at  the  beginning,  rapidly  in- 
crease in  number,  colleeting  at  the  margins  of  the  platelet  masses  and 
between  them.  Not  until  the  leucocytes  have  accumulated  does  the 
fibrin  appear.  As  Welch  remarks,  these  findings  afford  no  conclusive 
evidence  as  to  whether  fibrin-ferment  is  formed  from  the  leucocj^tes 
or  from  the  jilatelets,  but  since  the  fibrin  does  not  appear  until  after 
the  leucocytes  have  accumulated,  and  also  since  small  thrombi  may 
consist  solely  of  platelets  without  fibrin,  it  seems  probable  that  the 
leucocytes  must  be  looked  upon  as  the  chief  source  of  the  ferment. 
If  the  blood  is  made  ineoagulalile  by  injection  of  hirudin,  injury  to 
the  vessel-walls  causes  the  formation  of  thrombi  composed  entirely  of 
platelets  (Schwalbe).  Sometimes  small  clots  may  form  without  the 
apparent  ])articipation  of  either  platelets  or  leucocytes.  These  purely 
fibrinous  thrombi  seem  to  start  from  injured  endothelial  cells,  par- 
ticularly in  inflammatory  conditions,  such  as  pneumonic  lungs,  and 
give  the  impression  that  the  coagulin  is  derived  from  the  endothelial 
cells.  Zurhelle  attributes  by  far  the  most  important  part  to  the 
platelets,  an  opinion  shared  by  many,  including  Derewenko,^'''  who 
holds  that  the  coagulation  of  blood  with  entirely  occluded  vessels  is 
quite  distinct  from  true  thrombosis  because  of  the  lack  of  platelets 
in  stagnant  blood."  Clots  formed  in  the  absence  of  platelets  do  not 
shrink  like  proper  thrombi  fDuke). 

The  process  of  clotting  in  the  stoppage  of  hemorrhage  offers  some 
differences  from  intravascular  clotting,  in  that  the  coagulins  of  the 
tissue-cells  also  come  into  play.  It  is  rather  difficult  to  determine 
how  much  of  the  coagulation  depends  on  these,  and  how  much  on  the 
coagulins  of  the  leucocytes,  for  the  same  conditions  that  favor  libera- 
tion of  tissue  coagulins,  i.  e.,  much  laceration  and  destruction  of  the 
tissue,  also  favor  the  disintegration  of  leucocytes  by  offering  large 
areas  of  surface  for  contact.  Loeb  is  of  the  opinion,  however,  that 
of  the  two,  the  latter  factor  is  the  more  important.  It  may  be  re- 
called that  the  joint  action  of  tissue  and  blood  coagulins  is  greater 
than  the  sum  of  their  individual  actions,  which  also  must  be  an  im- 
portant factor  in  causing  clotting  in  bleeding  wounds. 

As  to  the  relative  importance  of  stagnation  and  vessel  injury  in 
producing  thrombosis,  we  know  that  total  stasis  in  an  uninjured  vessel 
may  not  result  in  thrombosis,  and,  on  the  other  hand,  extensive  in- 
jury or  large  calcified  plaques  in  the  intima  of  the  aorta  may  also 
cause  no  thrombosis  because  of  the  rapidity  of  the  blood  flow ;  and, 
furthermore,  clotting  may  occur  even  in  intact  vessels  under  the  influ- 
ence of  substances  liberating  fibrin-ferment  in  the  blood;  e.  g.,  snake 
venoms,  nucleoprotein  injections,  and  possibly  in  disease.  As  the  red 
corpuscles  contain  thromboplastic  substances  we  may  have  thrombi 
formed   when   hemolytic    agents    are   present   in    relatively   stagnant 

50  Ziegler's  Beitj-.,  1910   (48),  123. 
5"  Not  accepted  by  Schwalbe,  loc.  cit. 


324  DISTURBANCES    OF    CIRCULATION 

blood,  even  without  injury  to  the  vessel-walls."*  Presumably  the  clot- 
ting does  not  occur  when  the  stream  is  rapid,  because  any  fibrin- 
ferment  that  may  be  liberated  by  injured  leucocytes  or  endothelium 
is  swept  away  before  fibrin  can  become  attached  to  the  vessel-wall; 
or,  according  to  Howell's  hypothesis,  because  the  current  brings  an 
excess  of  antithrombin  to  the  point  where  the  thromboplastin  is  being 
formed.  Naturally,  the  combination  of  an  injured  vessel-wall,  a  slow 
current,  and  a  high  coagulability  offer  the  most  favorable  conditions, 
and  we  owe  to  Welch  the  appreciation  of  the  fact  that  in  a  large  pro- 
portion of  all  thrombi,  even  those  caused  by  apparently  purely  me- 
chanical agencies  (e.  g.,  cardiac  incompetence),  bacteria  are  present 
and  probably  determine  the  injury  to  the  vessel-walls  and  the  libera- 
tion of  fibrin-ferment.'^''  We  have  previously  referred  to  L.  Loeb's 
observations  on  the  effect  of  bacteria  in  causing  coagulation  of  the 
blood. 

Hyalin  thrombi  are  frequently  the  cause  of  extensive  degenerative 
lesions  in  the  viscera,  and  although  commonly  formed  of  red  corpuscles, 
they  do  not  stain  at  all  like  normal  corpuscles,  presumably  because 
a  certain  proportion  of  the  hemoglobin  has  been  altered  or  lost  through 
hemolysis.  Of  particular  interest  is  their  reaction  to  Weigert  's  fibrin 
stain,  by  which  they  often,  but  not  always,  stain  intensely:  a  fact  that 
has  been  the  cause  of  much  confusion  in  earlier  studies.  Flexner  '^^ 
first  appreciated  the  nature  of  these  thrombi  as  originating  from  ag- 
glutinated red  corpuscles,  although  Klebs,  Ziegler,  and  others  had 
earlier  suggested  that  hyalin  thrombi  were  formed  from  red  corpus- 
cles. Boxmeyer  "^  independently  arrived  at  the  same  conclusion  as 
Flexner,  in  studying  hyalin  thrombi  as  the  cause  of  necrosis  in  the 
liver  of  animals  infected  with  the  hog-cholera  bacillus.  Flexner  pro- 
duced hyalin  thrombi  by  injecting  eor]iuscles  agglutinated  by  ricin, 
or  by  injecting  ricin  itself,  or  hemolytic  substances  such  as  ether  or 
foreign  serum.  As  the  thrombi  become  old,  the  corpuscles  lose  their 
form  and  color  and  produce  the  typical  hyalin  appearance.  Pearce  "- 
proved  conclusively  the  dependence  of  the  thrombus  formation  upon 
agglutination,  for  he  secured  the  same  results,  including  the  liver  ne- 
crosis, by  injecting  specific  agglutinating  serums.  He  states  that  fib- 
nn  threads  may  occasionally  be  found  at  the  periphery  of  the  larger 
thrombi,  but  never  in  the  smaller  ones.  The  tendency  of  the  thrombi 
to  stain  like  fibrin  by  Weigert 's  method  is  observed  particularly  when 
the  tissues  have  been  hardened  in  Zenker's  solution.  It  is  extremely 
probable,  from  Flexner 's  observations,  that  in  the  thrombosis  pro- 
duced by  injecting  various  toxic  substances  into  the  blood,  the  so- 
bs Dietrich,  Cent.  f.  Path.   (Verhandl.) ,  1912   (23),  .372. 

53  Welch,  Venous  Thrombosis   in   Cardiac   Disease,  Trans.   Assoc.    Anier.    Phys., 
1900,  vol.  15. 

«oJour.  Med.  Research,  1002    (S),  .110. 

"Jour.  :Med.  Kesearcli,  ino:{   (0),  140. 

"2  Jour.  Med.  Research,  1!)04  (12),  329;  ih'uL,  1000  (14),  .-)41. 


EMIiOLISU  325 

called  " fih rin- ferment  ihromhosis,"  tlie  lliroinl)!  are  merely  agglu- 
tinative thrombi,  devoid  of  fibrin ;  this  is  undoubtedly  true  for  many 
of  the  thrombi  observed  after  poisoning  with  the  ]iowerfn1Iy  agglutin- 
ative snake  venoms  (see  Chap.  vi.).  Bacterial  hemagglutinins  may 
also  cause  the  formation  of  hyalin  thrombi.*'''  On  the  other  hand, 
some,  at  least,  of  the  hyalin  capillary  thrombi  are  undoubtedly  com- 
posed of  soft  masses  of  fibrin  which  have  not  become  fibrillar,  al- 
though the  successful  staining  by  fibrin  stain  is  not  final  proof  of  the 
fibrinous  nature  of  a  thrombus.  The  liver  necrosis  which  follows  ether 
injections  in  animals  is  caused  by  fibrinous  thrombi  which  result  from 
liberation  of  coagulins  by  the  injured  cells  (L.  Loeb). 

Secondary  Changes  in  Thrombi. — The  changes  that  occur  in 
thrombi  after  they  have  existed  for  some  time  are  largely  due  either 
to  ingroA^i;li  of  new  tissue  or  to  calcification,  the  latter  of  which  will 
be  considered  in  a  separate  chapter.  The  only  other  change  of  inter- 
est from  the  chemical  standpoint  is  the  central  softening  which  may 
occur  in  any  large  thrombus,  but  is  particularly  often  observed  in  the 
large  globular  thrombi  found  in  the  heart.  The  center  of  the  throm- 
bus may  be  so  completely  softened  that  it  resembles  a  sac  of  pus,  the 
contents,  according  to  Welch,  consisting  of  necrotic  fatty  leucocytes, 
albuminous  and  fatty  granules,  blood-pigment  and  altered  red  corpus- 
cles, and  occasionally  acicular  crystals  of  fatty  acids.  Undoubtedly 
this  softening  is  related  to  the  process  of  fibrinolysis  previously  de- 
scribed, and  depends  upon  digestion  of  the  fibrin  by  leucocytic  en- 
JijTnes,®*  but  the  fact  that  the  central  portion  alone  undergoes  soften- 
ing is  of  interest,  suggesting  that  the  antibodies  for  leucocytic  pro- 
teases, which  Opie  ^'^  found  present  in  normal  serum,  prevent  digestion 
at  the  surface  of  the  clot.  The  same  fact  indicates  that  the  tissue 
fibrinolysins  ^^^  do  not  plaj'  an  active  part  in  softening  clots. 

EMBOLISM 

Emboli  offer  little  of  chemical  interest,  because  of  the  purely  me- 
chanical nature  of  their  origin  and  of  the  effects  they  produce.®*^  An 
exception  exists  in  the  case  of  fat  emholism,  for  the  manner  in  which 
the  fat  is  removed  from  the  blood  has  invited  considerable  specula- 
tion."' Part  of  the  fat  is  eliminated  in  the  urine, "'^  but  the  problem 
of  how  it  escapes  from  the  glomerular  capillaries  is  not  satisfactorily 
explained ;  large  emboli  undoubtedly  lead  to  rupture  of  the  capillary 
walls,  and  probably  some  fat  also  escapes  through  stomata  or  similar 

63  Pearce  and  Wiiine,  Amer.  Jour.  Med.  Sci..  Oct.,  1904. 

64  Barker,  Jour.  Exp.  ]\[od.,  Ifl08    (10),  343. 

65  Jour.  Exper.  Med..  190.5    (7),  316. 

65a  See  Fleisher  and  Loeb.  Jour.  Biol.  Chem.,  1915   (21),  477. 

66  Fat  embolism  mav  follow  poisoning  with  potassium  chlorate  (Winosradow, 
Virchow's  Arch.,  1907  "( 190) .  92) . 

67  Full  discussion  by  Beneke,  Ziegler's  Beitr.,  1897   (22).  343. 

67a  Discussed  by  Sakaguchi,  (Biochem.  Zeit.,  1913  (48),  1)  who  finds  a  little 
fat  in  the  normal  urine. 


326  DISTURBAyCES    OF    CIRCULATION 

intercellular  openings.  Fat  may  also  escape  in  the  bile,  and  some  is 
probably  taken  up  by  the  tissue  and  endothelial  cells  by  phagocytosis. 
Beneke  found  that  the  fat  becomes  partly  emulsified  by  the  mechanical 
action  of  the  blood  current,  aided  to  a  slight  extent  by  saponification. 
The  larger  droplets  after  lodging  in  the  capillaries  are  surrounded 
by  leucocytes,  to  which  Beneke  ascribes  an  active  part  in  the  removal 
of  the  fat  as  fine  droplets  by  phagocytic  action.  We  may  well  believe, 
however,  that  the  lipase  of  the  plasma  is  an  important  agent  in  disin- 
tegrating the  emboli,  although  its  action  is  limited  because  of  the  rel- 
atively small  surface  which  the  large  drops  offer  for  attack.  After 
fat  droplets  have  been  taken  into  the  cells,  they  presumably  are  util- 
ized in  metabolism  like  normally  acquired  fat.  as  described  previously. 

The  amount  of  fat  free  in  the  blood  in  fat  embolism  may  be  sur- 
prisingly large.  Bissell  ^'^^  found  from  2  to  6.5  per  cent,  in  the  venous 
blood  of  several  typical  cases,  although  sometimes  figures  within  normal 
limits  (0.2  to  0.6  per  cent.)  were  found.  The  higher  quantities  repre- 
sent such  a  great  amount  of  free  fat  in  the  blood,  even  witliout  con- 
sidering the  quantity  held  in  the  capillaries,  that  it  is  scarcely  possible 
for  it  all  to  have  come  from  the  fractured  bones. 

Air  embolism  presents  some  features  of  interest  from  the  chemical 
standpoint,  especially  in  those  cases  following  sudden  decrease  in  at- 
mospheric pressure  in  persons  who  have  been  exposed  for  some  time  to 
pressures  considerably  higher  than  that  of  the  atmosphere  (diver's 
palsy,  caisson  disease,  etc.).  This  form  of  air  embolism  is  due  to  the 
fact  that  fluids  can  dissolve  much  more  gas  at  high  pressures  than  at 
low  pressures;  consequently  when  the  abnormally  great  pressure  to 
which  divers,  caisson  workers,  etc.,  are  subjected  is  too  suddenly  re- 
duced to  that  of  the  atmosphere,  the  excessive  gas  that  was  absorbed 
during  the  period  of  high  pressure  by  the  blood  and  tissue  fluids  is 
released,  and  forms  bubbles  in  the  tissues  and  blood.  The  bubbles  in 
the  nervous  tissues  may  cause  paralyses  of  various  sorts,  or  death; 
those  in  the  blood  may,  if  in  sufficient  amount,  cause  serious  or  fatal 
capillary  obstruction.  The  bubbles  consist  chiefly  of  nitrogen,  be- 
cause the  power  of  the  blood  to  hold  oxygen  in  combination  is  so  great 
that  not  much  of  this  gas  becomes  freed.'^*  The  body  fluids  of  normal 
persons  contain  about  675  c.c.  of  nitrogen,  all  told,  but  at  22  pounds 
pressure  this  is  increased  to  1350  c.c,  while  but  about  50  c.c.  of  free 
oxygen  would  be  present  (Langlois).  Carbon  dioxide  is  so  readily 
combined  in  the  blood  that  none  is  free  even  at  high  pressure,  al- 
though ]\IcWhorter ""  reports  that  the  gas  collected  from  the  right  side 
of  the  heart  in  a  fatal  case  contained  20  per  cent.  CO,  and  80  per  cent, 
nitrogen.     Possibly  some  oxygen  may  also  be  released  from  solution 

iTb.Tour.  Amer.  Med.  Assoc,  IIUC)   (Cu),  l!>-2<>. 

ns  Tliis  subject  is  fully  discussed  by  Leonard  Hill  in  ■rvccciit  Advances  in 
Physiolof^v  and  Biocliemistrv,"  T»ndon,  190G. 

eoAmer.  Jour.  Med.  Sci., 'lOlO  (130),  373;  Erdman.  ibid.,  V^\^^    (145).  friO. 


jyFAncTiox  327 

(luring-  dec()uii)ivssion.'"  At  body  temperature  fats  can  dissolve  five 
times  as  much  nitrogen  as  serum  or  plasma/^  which  probably  accounts 
for  the  severity  of  the  changes  in  the  nervous  system  with  its  rich 
lipoid  content  and  delicate  structure.  Air  embolism  following  obstet- 
rical operations  oi-  surgical  operations  about  the  neck  and  chest  pre- 
sents chiefly  mechanical  features,'-  and  large  (luantities  of  air  nuist  be 
present  to  cause  dangerous  obstruction  to  circuhition.'^  Gas-bubbles 
may  be  produced  in  the  blood  soon  after  death  by  B.  aerogenes  cap- 
sulatus,  but  it  is  not  probable  that  they  are  formed  before  death  and 
cause  air  embolism. 

INFARCTION 

The  changes  that  occur  in  infarcted.  areas  are  of  much  interest  in 
connection  with  the  study  of  autolysis,  for  the  absoi^ption  of  the  ne- 
crotic tissue  of  aseptic  infarcts  is  purely  a  matter  of  autolysis.  Ja- 
coby  '*  found  by  ligating  off  a  portion  of  a  dog's  liver,  and  keeping  the 
dog  alive  for  some  time  afterward,  that  in  the  infarcted  tissues  so 
produced  leucine  and  tyrosine  could  be  detected,  just  as  they  are 
found  in  liver  tissue  undergoing  autolysis  outside  of  the  bodj'.  So, 
too,  proteoses  may  appear  in  the  urine  when  any  considerable  amount 
of  tissue  is  cut  off  from  its  blood-supply.  The  processes  of  autolysis 
which  occur  in  ordinary  sterile  infarcts  are,  however,  extremely  slow, 
and  it  is  doubtful  if  enough  of  the  products  are  ever  in  the  blood  or 
urine  at  any  one  time  to  be  detected  or  to  cause  noticeable  effects. 
For  example,  in  an  infarct  of  the  kidney  which  was  known  to  be  al- 
most exactly  fourteen  weeks  old,  there  still  remained  a  la.yer  of  ne- 
crotic cortex  one  millimeter  thick,  quite  unabsorbed  during  this  time. 
If  we  examine  such  aseptic  infarcts  in  various  stages,  we  get  the  im- 
pression that  the  digestion  is  accomplished  by  leucocytes  acting  on  the 
j;eri])hery  of  the  infarct,  and  not  entering  the  dead  area  deeply,  pre- 
sumably because  of  a  lack  of  cheraotactic  substances  in  the  dead  cells. 
On  the  other  hand,  it  seems  probable  that  the  tissue  enzymes  them- 
selves play  an  important  part  in  the  autolysis,  for  if  we  implant  into 
inimals  pieces  of  tissue  in  which  the  enzymes  have  been  destroyed  by 
heating  to  boiling,  it  will  be  found  that  the  cells  and  their  nuclei  re- 
main unaffected  for  man}'  weeks;  whereas  if  sterile  unheated  pieces 
of  tissue  in  which  the  enzymes  are  still  active  are  implanted,  they  lose 
their  nuclear  stain  and  begin  to  disintegrate  relatively  soon,  without 
apparent  participation  by  the  leucocytes."''  Ribbert  '*'  found  that  in 
experimentally  produced  anemic  infarcts  in  the  kidneys  of  rabbits  the 
nuclei  retain  their  staining  property  well  for  nearly  twenty-four  hours^ 

-oHill  and  Greenwood,  Proc.  Roval  Soc.    (B).  1907    (79),  284. 

71  Vernon,  i1>id.,  p.  Sfifi:  Quincke",  Arcli    exp.  Path.  u.  Pharni..  1910  (02).  404. 

"2  Review  of  literature  by  Wolff,  Virchow'.s  Archiv.,  190,3    (174).  454. 

T3  See  Hare,  Amer.  Jour.' :\rod.  Sciences,  1902    (124),  84:5. 

■^Zeit.  phvsiol.  Chem.,  1900    (.30).  149. 

T3  Wells,  Jour.  :Med.  Research.  1906   (15),  149. 

ToVirchow's  Arch.,  1899   (155),  201. 


328  DISTURBAXCES    OF    CIRCULATIOy 

becoming  tlieii  small  and  deeply  stained,  undergoing  karyorrhexis^ 
and  in  large  part  disappearing  from  the  convoluted  tubules  inside  of 
forty-eight  hours,  although  some  nuclei  may  persist  in  the  glomerules- 
for  three  or  more  days.  In  human  infarcts,  Rihbert  believes,  the 
process  goes  on  faster,  for  he  has  observed  here  a  loss  of  nuclei  within 
twenty-four  hours.  These  nuclear  changes  undoubtedly  depend  upon, 
autolysis,  but  it  is  probable  that  the  enzymes  concerned  reside  in  the 
cytoplasm  rather  than  in  tlie  nucleus,  for  I  have  observed  that  cells 
of  lymphoid  type,  with  practically  no  cytoplasm,  generally  retain 
their  nuclear  stain  much  longer  than  cells  with  more  cytoplasm;  this- 
is  particularly  noticeable  in  splenic  infarcts,  where  the  ]\Ialpighian 
corpuscles  retain  their  staining  affinities  much  longer  than  the  pulp 
elements.  AVhether  the  destruction  of  the  nuclei  is  accomplished 
by  the  ordinary  intracellular  proteases,  or  by  special  nucleoprotein- 
splitting  enzymes  (nuclease, ^^  etc.),  remains  to  be  determined.  It  is 
quite  possible,  however,  that  the  first  changes  consist  of  a  splitting 
of  the  nucleoproteins  of  the  nucleus  by  the  autolytic  enzymes,  liber- 
ating the  nucleic  acid,  which  gives  the  nuclei  the  characteristic  intense- 
staining  with  basic  dyes  (pycnosis)  observed  in  areas  of  early  anemic 
necrosis.  The  nucleic  acid  may  then  be  further  decomposed  by  the 
nuclease  or  similar  enzymes.  Taken  all  together,  then,  it  would  seem 
that  the  nuclear  and  cellular  alterations  that  make  up  the  character- 
istic picture  of  anemic  necrosis  are  brought  about  by  the  intracellular 
enzj^mes — an  autolytic  process.  The  removal  of  the  dead  substance, 
however,  seems  to  be  accomplished  rather  by  the  invading  leucocytes, 
through  heterolysis.  The  relatively  small  part  taken  by  the  intracel- 
lular enzymes  may  possibly  be  due  to  the  seeping  through  them  of  alka- 
line blood-plasma,  for  autolytic  enzymes  are  not  active  in  an  alkaline 
medium;  the  leucocytic  enzymes,  however,  act  best  in  an  alkaline 
medium.'^^ 

About  the  periphery  of  infarcts  is  usually  observed  more  or  less  fat 
deposition  (Fischler),'^''  particularly  in  the  endothelial  cells  (Ribbert). 
This  is  not  peculiar  to  infarcts,  however,  for  Sata  ^"  found  a  similar 
peripheral  fatty  metamorphosis  common  to  all  necrotic  areas.  The 
basis  of  this  is  possibly  the  persistence  of  the  cell  lipase,  which  syn- 
thesizes fatty  acid  and  glycerol  dififusing  into  the  necrotic  area  with 
the  plasma,  unchecked  by  the  normal  oxidative  destruction  of  these 
substances.      (See  "Fatty  Degeneration,"  Chap,  xiv.) 

Hemorrhagic  infarcts  offer,  in  addition  to  the  changes  common 
to  anemic  infarcts,  the  alterations  ocQurring  in  the  blood-corpuscles. 
Panski  ^^  found  that  after  ligation  of  the  splenic  vein  of  dogs  the  red 

T7  Sachs,  Zoit.  physiol.  Cliem.,  lOUu   (4C),  337;  Schiltonliclm.  ibid..  3.")4. 

78  More  fullv  discusapd  bv  WoUs.ts  Joe.  cit.,  and  under  necrosis.  Cluip.  xiii. 

-"Cent.  f.  Patlt.,  1002    (13),  417. 

«oZiepler'8  Beitr.,  1000   (2R),  4()1. 

SI  "Untcrsucliuii^^-n   iilicr  den   I'i^niieiil j^clialt  der   Stamingsmilz,"  Dor|)at.    1800. 


INFAUCTIOX  329 

corpuscles  begin  to  give  up  their  heuioglobiu  in  about  tliree  hours. 
After  twelve  hours  fibrin  formation  begins  in  the  tissues,  the  corpus- 
cles continue  to  give  up  hemoglobin  and  become  cloudy  in  appearance. 
Later,  iron-containing  pigment  is  formed  in  the  cells  beneatli  the  cap- 
sule, but  in  the  deeper  tissue  even  the  iron  normally  present  in  the 
spleen  tissue  seems  to  disappear;®-  this  possibly  depends  upon  the 
fact  that  pigment  reacting  for  iron,  hemosiderin,  is  formed  only  in 
living  colls  under  the  influence  of  oxygen,  or  it  may  be  that  acids 
formed  during  autolysis  dissolve  it.  During  autolysis  iji  vitro,  how- 
ever, Corper  "*•'  found  no  evidence  of  removal  of  iron  from  insoluble 
or  coagulable  compounds.  The  hemolysis  is  probably  produced  either 
by  the  action  of  autolytic  products,  which  are  notoriously  hemolytic, 
or  perhaps  also  by  direct  attack  of  tissue  and  blood  proteases  upon 
the  corpuscles. 

Other  retrogressive  changes  that  may  occur  in  infarcts,  such  as  sep- 
tic softening  and  calcification,  are  not  greatly  different  from  the  same 
processes  occurring  in  other  conditions,  and  will  be  considered  with 
the  discussion  of  these  processes. 

s2See  also  M.  B.  Schmidt,  Cent.  f.  Path.,  1908   (19),  416. 
S3  Jour.  Exper.  Med.,  1912    (15),  429. 


CHAPTER   XII 

EDEMA  1 

As  the  term  edema  indicates  the  excessive  accumuhitioii  of  lymph 
(which  may  be  either  normal  or  modified  in  composition )  in  the  cells, 
intercellular  spaces,  or  serous  cavities  of  the  body,  the  problems  of 
edema  are  inseparably  connected  Avith  the  consideration  of  the  proc- 
esses of  physiological  formation  and  removal  of  lymph.  For  many, 
years  the  study  of  these  processes  has  been  a  favorite  field  of  investi- 
gation by  physiologists,  and  the  great  battle-place  of  the  "vitalistic" 
and  "mechanistic"  schools;  and  to  this  day  the  forces  that  determine 
the  formation  of  lymph  and  its  subsequent  absorption  have  not  been 
completely  understood.  By  the  application  of  the  principles  of  phys- 
ical chemistry  to  the  problem,  however,  great  advances  have  recently 
])een  made,  which  seem  to  render  our  understanding  of  both  lymph- 
formation  and  its  pathological  accumulation  in  the  tissues  much 
clearer  and  more  nearly  accurate  than  they  were  before.  AVe  shall 
first  consider,  therefore,  the  physiological  formation  of  lymph,  before 
taking  up  the  subject  of  edema. 

Composition  of  Lymph. — Lymph  consists  of  material  derived  from  two  chief 
sources.  The  greater  part  consists  of  fluid  passing  out  of  tlie  capiUaries  into 
the  tissue  spaces:  here  it  is  modified  by  the  addition  of  products  of  metabolism 
derived  from  the  tissue-cells,  and  by  the  sul)traction  of  materials  tliat  the  cells 
utilize  in  their  metabolism.  It  is,  therefore,  essentially  a  modified  blood  plasma, 
and  the  modifications  the  plasma  undergoes  are  so  slight  tliat.  under  ordinary 
conditions,  lymph  shows  on  analysis  no  considerable  differences  from  blood 
plasma,  except  a  relative  poverty  in  proteins,  due  chiefly  to  the  impermeability 
of  the  capillary  walls  for  colloids.  Its  quantitative  composition  varies  greatly, 
depending  upon  the  conditions  under  which  it  is  collected,  whether  during 
activity  or  rest,  etc.  Tiie  following  tables  of  analyses  have  been  collected  by 
Ilammarsten: 

1  2  .*?  4 

Water 0.30.0  0.34.S  057.6  0.55.4 

Solids         00.1  05.2  42.4  44.6 

0.4  2.2 


Fibrin         

0.5 

0.0 

Albumin 

42.7 

42.8 

Fat,  Cholesterol,  Lecithin  . 

.S.S 

0.2 

Extractive    bodies     . 

5.7 

4.4 

Salts 

7.3 

S.2 

.34.7 


35.0 


1  and  2  are  analyses  of  lymph  from  tlie  tliigli  of  a  woman.  .'!  is  from  tlic 
contents  of  sac-like  dilated  vessels  of  the  spermatic  cord,  4  is  lymph  from  the 
neck  of  a  colt. 

lA  complete  bibliograpliy  is  given  by  Afeltzer.  .\merican  l\redicine,  1004  (S), 
10  et  seq. ;  also  bv  Klemeiisiewicz,  in  Krehl  and  Marchand's  Ihuullmeli  d.  allg. 
Path.,  1012,  II  (1),  341:  ]\Iagnus.  TTandlmeh  d.  Hiochem..  1!)(1S,  11  |2),  90; 
Gorhartz,  ihid.,  ]>.  11(5. 

330 


Foinf.iTJox  or  L)  Mi'if  331 

Cliyle  difl'ers  from  lym]ili  chit'lly  in  tlie  prosem-t'  of  larjic  (juantities  of  fat; 
(hiring  starvatiim  the  lymj)li  and  tiie  t-liyle  arc  of  jiractically  tlie  same  composi- 
tion. 

Normal  lymph  contains  much  less  fibrinogen  tlian  docs  tlic  hlocvd  plasma,  and 
hence  coagulates  slowly.  Lipase  and  oilier  enzymes  ha\-e  lieen  found  in  the 
lymph,  as  in  the  plasma.  'I'lie  products  of  tissue  metabolism  added  to  the 
lymph  by  the  cells  may  render  it  toxic  (Asher  and  Barbera^).  I'nder  patii- 
ological  conditions  llie  lym])h  may  l)e  greatly  altered,  liecoming  jxwrer  in  solids 
under  some  conditions  of  edema,  and  bee  nning  rich  in  proteins  and  l)loo(l-cor- 
puscles  under  intlammatory  conditions,  luitil  it  ])artakes  of  tlie  characteristics  of 
an  inflammator}-  exudate   (see  analyses  of  transudates  and  exudates). 

An  important  fact  to  consider  is,  that  of  the  entire  water  of  the 
body  hut  about  one-tenth  is  in  the  blood.  About  two-thirds  of  the 
entire  weight  of  the  body  is  water,  which  is  mostly  in  the  cells  and 
tissues,  firmly  bound  by  the  colloids,  only  an  unknown  but  smaller 
portion  being  as  free  movable  fluid,  and  even  here  always  associated 
with  more  or  less  colloid.  A  body  weighing  60  kilos  will,  therefore, 
have  40  kilos  of  water,  of  which  but  about  4  kilos  is  blood. 

FORMATION  OF  LYMPHS 

Filtration  Theory. — The  simplest  possible  conception  of  lymph 
foruuition  is  that  it  is  merely  the  result  of  filtration  of  the  liquid  con- 
stituents of  the  blood  through  the  capillary  walls  under  the  influence 
of  the  blood  pressure.  This  "filtration  theory"  was  supported  origi- 
nally by  Ludwig,  and  it  w^as  a  prominent  factor  in  the  early  appli- 
cations of  mechanical  principles  to  biological  processes.  In  support 
of  this  theory  were  advanced  the  results  of  numerous  experiments  in 
which  it  was  shown  that  increasing  the  blood  pressure  by  means  of 
ligating  the  veins,  or  by  causing  arterial  dilatation,  resulted  in  an  in- 
crease of  the  lymph  flowing  out  of  the  lymph-vessels  of  the  part. 
Also,  when  the  blood  pressure  is  raised  by  epinephrine  or  by  other 
means,  a  large  proportion  of  the  fluid  leaves  the  blood  vessels;  con- 
versely, M^hen  the  blood  pressure  is  suddenly  lowered  by  hemorrhage 
there  is  a  rapid  passage  of  fluid  from  the  tissues  into  the  blood.  The 
experimental  results  were  not  always  favorable  to  the  theory,  how- 
ever, particularly  in  the  experiments  in  which  blood  pressure  was 
raised  by  arterial  dilatation ;  often  the  flow  of  lymph  was  little  in- 
creased, even  w^hen  the  arterial  flow  and  pressure  were  greatly  in- 
creased. Nevertheless,  the  filtration  theory  held  for  many  years,  not 
only  as  an  explanation  of  lymph  formation,  but  also  as  an  explanation 
of  urinary  secretion  and  of  the  secretion  by  other  organs.  It  w^as 
only  within  a  comparatively  short  time  that  it  became  clear  that  filtra- 
t^'on  alone  ccmld  not  account  for  all  the  phenomena  of  secretion.  For 
example,  in  many  lower  forms  with  undeveloped  circulatory  systems, 
and  almost  no  blood  pressure,  secretion  goes  on  vigorously ;  the  pres- 
sure  of   glandular  secretions   may  be   much   higher  than   the   blood 

2Zeit.   f.   Biol..   1808    (36),    1.54. 

3  See  review  by  Asher,  Biochem.  Centralblalt.  1005   (4),  1. 


332  EDEMA 

pressure  within  the  capillaries ;  the  activity  of  secretion  is  by  no  means 
in  proportion  to  blood  pressure,  etc.  If  in  *^landular'  secretion,  there- 
fore, fluids  are  removed  from  the  blood  and  transferred  into  an  ex- 
cretory duct  through  the  action  of  some  force  other  than  that  of  the 
blood  pressure,  it  is  probable  that  lym])h  formation  is  eciually 
independent  of  blood  pressure.  On  this  basis  Ileidenliain  advanced 
his — 

Secretory  theory  of  lymph  formation,  in  which  he  suggested  that 
lymph  is  the  product  of  an  active  secretion  by  the  endothelial  cells  of 
the  capillaries,  just  as  saliva  is  the  product  of  the  activity  of  the 
glandular  cells.  He  showed  that  certain  chemical  substances  may 
stimulate  lymph  flow,  independent  of  blood  pressure,  just  as  pilocar- 
pine and  other  drugs  may  stimulate  the  secretion  of  saliva.  These 
hmiph-stimulating  substances,  which  he  named  lyinphagogues,  fall  into 
two  distinct  classes.  One  which  includes  such  substances  as  peptone, 
leech  extract,  strawberry  juice,  extracts  of  crayfish,  mussel  or  oysters, 
and  numerous  other  tissue  extracts,  are  characterized  by  causing  the 
secretion  of  a  lymph  which  is  rich  in  proteins,  even  richer  in  proteins 
than  the  blood  plasma;  and,  furthermore,  there  is  no  simultaneous 
increase  in  urinary  secretion.  Heidenhain  considered  that  these  sub- 
stances caused  lymph  secretion  by  stimulating  the  capillary  endothe- 
lium in  a  specific  manner ;  as  they  caused  no  appreciable  rise  in  blood 
pressure  the  increased  lymph  secretion  certainly  could  not  be  attrib- 
uted to  filtration.  This  independence  of  the  lymph  flow  of  blood 
pressure  is  most  conclusively  shown  by  postmortem  lymph  secretion; 
for  example,  Mendel  and  Hooker  *  observed  lymph  flow  for  four  hours 
after  death,  in  a  dog  that  had  received  an  injection  of  peptone  eight 
minutes  before  being  killed.^ 

The  second  class  of  lymphag-ogues  includes  crystalloidal  substances, 
such  as  sugar,  urea,  and  salts. ^'^  Lymph  secreted  under  the  influence 
of  these  substances  is  poorer  in  protein  than  ordinary  lymph,  and  at 
the  same  time  an  increased  urinary  secretion  is  produced.  With 
these  crystalloidal  lymphagogues  the  amount  of  effect  is  in  inverse 
proportion  to  their  molecular  M^eight,  which  means  that  their  effects 
depend  upon  the  number  of  molecules  in  solution  rather  than  upon 
their  nature ;  in  other  words,  the  stimulation  of  lymph  by  crystalloids 
is  dependent  upon  the  osmotic  pressure  of  the  crystalloids.  Heiden- 
hain explained  their  action  as  follows:  The  crystalloids  are  secreted 
into  the  lymph-spaces  by  the  action  of  the  capillary  endothelium,  and 
there,  owing  to  their  raising  osmotic  pressure,  cause  a  flowing  of 
water  out  of  the  vessels.     The  difificulfy  here  is  to  explain  why  the 

4Anier.  Jour,  of  Physiol.,  1902   (7),  380. 

OA  fact  not  sufTiciently  takon  into  account  is  that  blisters  filled  witli  scrum, 
i.  e.,  an  inflanimatorv  odoma,  mav  be  ])rodticcd  in  dead  bodies  bv  burns  or  scalds. 
(See  Leers  and  Kaysky.  Virclio\v''s  Arch.,  IDOO    (197),  .324). 

5a  The  action  of  many  other  substances  lias  In-en  invest ipatcnl  by  Vanaj^awa, 
Jour.  Pharmacol.,  1916  "(9),  75. 


FORMATION  OF  JA Wff'H  333 

crystalloids  while  still  in  the  vessels  do  not  attract  the  fluids  from  the 
lymph-spaces  into  the  blood,  and  so  canse  rather  a  lessened  lymph 
secretion. 

"Wliile  admitting-  that  in  pathological  conditions  (e.  g.,  passive  con- 
j^estion)  pressure  and  filtration  may  play  an  important  part,  Heiden- 
haiu  considered  that  an  active  secretion  by  the  endothelial  cells  is  the 
chief  factor  in  the  normal  formation  of  lymph.  The  means  by  which 
the  cells  perform  this  function  was  unknown ;  it  was  considered  as  an 
example  of  "vital  activity,"  Ileidenhain  meaning  by  this  term  such 
chemical  and  physical  forces  of  living  cells  as  are  unliJiown  or  not 
understood  at  the  present  time,  rather  than  any  metaphysical  concep- 
tion of  living  matter,  such  as  many  vitalists  assume. 

Other  observers,  corroborating  Heidenhain's  results  for  the  most 
part,  liave  modified,  or  amplified  his  theory.  Asher  and  his  collabo- 
rators, for  example,  ascribe  the  work  done  in  causing  lymph  forma- 
tion to  the  cells  of  the  various  tissues  and  organs,  rather  than  to  those 
of  the  capillary  wall.  The  increased  flow  of  lymph  from  the  salivary 
gland  that  occurs  during  its  activity  they  consider  due  to  the  work 
of  the  gland  cells,  and  its  function  the  removal  of  products  of  metab- 
olism. The  action  of  such  a  lymphagogue  as  peptone  they  ascribe  to 
its  stimulation  of  cellular  activity,  particularly  in  the  liver,  where  it 
causes  an  increased  formation  of  bile.  Gies  *'  and  Asher  also  ob- 
served that  after  an  injection  of  crystalloidal  lymphagogues,  such  as 
sugar,  a  prolonged  flow  of  lymph  occurred  after  the  death  of  the 
animal,  proving  completely  that  such  lymphagogic  action  is  inde- 
pendent of  blood  pressure. 

Potocytosis. — In  explanation  of  the  process  l)y  which  the  cells,  whether  en- 
dothelial or  tissue-cells,  pass  fluids  throuph  themselves  from  one  place  to  another, 
Meltzer  1  has  made  an  interesting  suggestion,  as  follows:  Considering  the  prop- 
erty of  endothelial  cells  to  act  as  phagocytes,  MacCallum  "  has  shown  tliat  solid 
granules  (e.  f/.,  coal  pigment,  carmin)  are  taken  throujrh  the  walls  of  the  lymphat- 
ics by  the  phagocytic  activity  of  their  endothelial  cells.  ]\Ieltzer  suggests  tliat  in 
a  similar  way  the  endothelial  cells  may  transport  through  the  vessel-walls  not 
only  solid  particles,  ])ut  also,  by  the  same  mechanism,  substances  in  solution; 
and  for  this  hypothetical  process  he  suggests  tlie  name  "potoci/tosis."  There  can 
be  little  question  that  cells  do  take  up  substances  in  solution,  and  sometimes  this 
is  done  in  an  apparently  selective  manner;  e.  g.,  the  taking  up  of  bacterial  toxins 
and  vegetable  poisons  in  the  peritoneal  cavity  by  the  leucocytes.  Presumably 
the  mechanism  of  "potocytosis"  is  not  different  from  that  of  phagocytosis,  chemo- 
tactic  forces  determining  the  occurrence  of  tlie  process.  Xo  experimental  evi- 
dence has  been  advanced  as  yet  for  this  very  plausible  hypothesis. 

Permeability  of  Capillaries. — In  explanation  of  the  variability 
in  the  amount  and  composition  of  the  lymph.  Starling  ^  has  introduced 
the  factor  of  altered  permeability  of  the  capillary  walls,  which  pre- 
sumably depends  upon  the  number  and  size  of  the  pores.  He  found 
that  normally  the  lymph  coming  from  the  lower  extremities  contains 

sAmer.  Jour.  Physiol..  IflOO   (.3),  p.  xix:  Zeit.  f.  Biol..  1000   UO),  207. 

7  Johns  Hopkins  Hosp.  Bull.,  1903   (14),  1 

s  Lancet,  1896   (i),  Jlay  9,  et  seq.;  Sch:i,fer's  Text-book  of  Physiology,  vol.  1. 


334  EDEMA 

only  2  per  cent,  to  3  per  eent.  of  proteins,  while  lymph  from  the  intes- 
tines contains  4  per  eent.  to  6  per  eent.,  and  lymph  from  the  liver  eon- 
tains  6  per  cent,  to  8  per  eent.  of  proteins ;  hence  he  considers  that  the 
liver  capillaries  are  the  most  permeable,  i.  e.,  have  the  largest  pores, 
so  that  more  of  the  laroe  colloid  molecules  can  escape  from  them.  The 
effect  of  lymphafi'oo'ues  of  the  first  class  (peptones,  etc.)  he  attributes 
to  their  poisonous  properties,  and  the  consetiuent  injury  to,  and  alter- 
ations in,  the  capillary  wall.  The  crystalloidal  lymphagogues,  he 
believes,  act  by  first  attracting  fluids  from  the  tissues  into  the  blood 
with  a  resulting  ''hydremic  plethora,"  which  in  turn  leads  to  in- 
creased blood  pressure  and  consequent  filtration  of  a  watery  fluid 
out  of  the  vessels.  He  considers,  therefore,  that  the  amount  and 
qii_ality  of  the  IxiBph  produced  in  any  part  are  determined  solely  by 
two  factors,  the  intracapillary;J)lood_:pjceasm:e  and  the  permeability  ^f_ 
the  capillary  wjiJjs. 

In  connection  with  this  question  of  the  permeability  of  the  capil- 
lary walls,  Meltzer  suggests  that  the  contractility  and  irritability  of 
the  endothelium  may  be  a  potent  factor  in  determining  the  size  of  the 
pores  in  the  capillary  walls.  When  in  a  tonic  condition,  the  endothe- 
lium is  firmly  contracted  about  the  pores,  keeping  their  size  small ; 
when  the  endothelial  cells  become  relaxed  by  any  cause,  such  as  poi- 
sons, high  blood  pressure,  poor  nourishment,  etc.,  the  pores  are  en- 
larged, and  increased  escape  of  fluids  results.  It  must  be  borne  in 
mind,  however,  that  most  histologists  do  not  now  admit  that  capillary 
walls  contain  pores. 

M.  H.  Fischer  holds  that  the  endothelial  cells  undergo  changes  in 
consistency  through  changes  in  the  affinity  of  the  cell  colloids  for  wa- 
ter; especially  under  the  influence  of  acids  the  endothelium  may  be- 
come much  more  fluid  and  of  greater  permeability.  Adolf  Oswald  " 
says  that  the  normal  capillary  wall  is  somewhat  permeable  for  the 
less  viscous  blood  proteins  (albumin  and  pseudoglobulin),  and  in  in- 
flammation this  permeability  becomes  increased  so  that  the  more  vis- 
cous euglobulin  and  fibrinogen  can  pass  through. 

Osmotic  Pressure. — Still  another  possible  factor  in  causing  fluid 
to  leave  the  vessels  is  osmotic  pressure.  Heidenhain  refers  to  this 
cause  the  transudation  produced  by  crystalloid  lymphagogues,  al- 
though in  a  rather  unsatisfactory  manner.  As  a  result  of  the  more 
recent  studies  of  physical  chemistry,  and  its  application  to  biological 
processes,  vce  have  learned  to  appreciate  the  importance  of  osmotic 
pressure  in  cell  activities  (see  Introductory  Chapter),  and  in  the 
question  of  lymph  formation  it  occupies  a  ])articularly  important 
])lace.  AVe  may  consider  it  as  follows:  In  the  blood  we  have  certain 
proportions  of  readily  diffusible  crystalloids  and  of  non-diffusible 
colloids.  If  no  metabolic  processes  were  going  on  in  the  tissues,  we 
should  have  the  diffusible  substances  leaving  the  vessel-walls  (leaving 

»  Zcit.  f.  oxp.  Patli..  1010   (S),  22(). 


FORM  AT/OX   OF  JAM  I'll  335 

out,  for  tile  j)rc.sent,  any  question  of  activity  on  the  part  of  the  endo- 
Tlielium)  until  an  osmotic  equilibrium  is  established  in  the  tissues  and 
in  the  blood.  As  a  matter  of  fact,  however,  the  blood  proteins  are  not 
absolutely  non-diifusible,  but  small  quantities  do  pass  through  the  cap- 
illary walls,  and  so  lymph  under  such  a  hypothetical  condition  would 
consist  of  a  mixture  of  the  same  osmotic  concentration  as  the  blood 
plasma,  with  about  the  same  proportion  of  crystalloids,  but  a  smaller 
proportion  of  proteins ;  this,  it  will  be  noticed,  is  just  about  the  com- 
position of  normal  lymph.  During  life,  however,  the  cells  of  the  tis- 
sues are  causing  metabolic  changes  in  these  lym])hatic  constituents, 
and  these  changes  consist  chiefly  in  breaking  down  large  molecules  of 
proteins,  carbohydrates,  and  fats  into  much  smaller  molecules.  Now 
the  osmotic  pressure  of  a  solution  is  dependent  upon  the  numher  of 
molecules  and  ions  it  contains,  hence  by  breaking  down  these  few 
large  molecules  with  verj^  little  osmotic  pressure  into  many  small  mol- 
ecules, the  osmotic  pressure  in  these  cells  and  tissues  becomes  raised 
above  that  of  the  blood-vessels,  and  consequently  water  flows  out  of 
the  vessels  because  of  the  increased  pressure.  We  see  here  the  prob- 
able explanation  of  the  stimulating  influence  of  metabolic  products 
upon  the  formation  of  lymph,  noted  by  Hamburger,  Heidenhain,  and 
others.  For  suggesting  and  urging  the  importance  of  osmotic  pres- 
sure in  the  formation  of  lymph  we  are  indebted  particularly  to  Hei- 
denhain, V.  Koranyi,^"  J.  Loeb,^^  and  Roth.^-  Loeb  show^s  very  clearly 
the  relative  greatness  of  the  water-driving  force  of  osmotic  pressure 
as  compared  to  that  of  blood-pressure,  by  his  statement  that  the  os- 
motic pressure  of  a  physiological  salt  solution  is  about  4.9  atmospheres, 
which  is  twenty  ti)nes  as  great  as  the  hJood  jjressure  with  which  we 
have  to  do  in  ordinary  physiological  experiments.  In  varying  osmotic 
conditions  we  may  readily  see  an  explanation  for  the  increased  lymph 
flow  that  occurs  during  tissue  activity ;  namely,  it  is  due  to  the  in- 
creased formation  of  metabolic  products.  ]Many  of  the  lymphagogues 
may  act  similarly  by  stimulating  metabolic  activity,  with  resulting  in- 
crease in  the  formation  of  osmotic  pressure-raising  products  of  metab- 
olism in  the  organs ;  e.  g.,  the  increased  lymph  flow  from  the  thoracic 
duct  that  follows  stimulation  of  hepatic  activity  by  injection  of  pep- 
tone (Heidenhain)  or  ammonium  tartrate  (Asher  and  Busch).^^  As 
we  shall  see  later  in  considering  edema,  osmotic  pressure  may  play  an 
important  part  in  the  pathological  formation  of  lymph.  It  must  be 
admitted,  however,  that  there  are  many  difficulties  in  the  way  of 
accepting  unqualifiedly  the  original  views  as  to  the  importance  of 
osmotic  pressure  in  lymph  formation.  For  example,  the  lymph  con- 
tains more  chlorides  and  may  have  a  much  higher  osmotic  pressure 

loZeit.  f.  klin.  Med..  1807    (33),  1:    1898    (34),  1. 
iiPfliicrer's  Arch.,   1898    (71),  457. 
12  Englemann's  Arch.,  1899,  p.  416. 
i3Zeit.  f.  Biol.,  190O   (40),  333. 


336  EDEMA 

than  the  serum  of  the  same  animal  (Hamburger,  Carlson,  et  al.) }^^ 
Variable  Capacity  of  Colloids  for  Water. — Colloids  of  the  type  of  the 
tissue  proteins,  i.  c,  liydrophil  colloids,  imbibe  water  with  great  avid- 
ity,  until  a  certain  proportion  of  water  is  present,  the  proportion 
varying  under  different  conditions.  The  importance  of  tliis  force  in 
the  production  of  edema  and  related  processes  was  first  pointed  out 
by  Martin  H.  Fischer,  and  has  been  developed  extensively  by  him." 
The  amount  of  water  which  a  given  hydrophil  colloid,  such,  for  exam- 
ple, as  gelatin,  or  fibrin,  will  take  up,  is  greatly  modified  by  the  reac- 
tion of  the  solution  and  by  its  content  of  electrolytes.  Very  small 
concentrations  of  acids  or  alkalies  will  greatly  increase  the  amount 
of  water  absorbed,  while  salts  reduce  it,  and  the  different  acids  and 
salts  vary  in  their  effects;  thus  hydrochloric  acid  causes  a  greater 
swelling  of  colloids  than  a  corresponding  strength  of  sulphuric  acid, 
and  calcium  chloride  depresses  the  swelling  more  than  potassium 
chloride.  The  effect  of  the  salts  is  made  up  of  their  constituent  ions. 
Non-electrolytes  have  relatively  little  effect.  The  forces  developed 
by  this  affinity  of  colloids  for  water  are  enormous ;  thus,  to  prevent 
the  taking  up  of  water  by  starch  requires  a  pressure  of  over  2500  at- 
mospheres, and  dried  gelatin  will  take  up  25  times  its  weight  of  water, 
and  fibrin  as  much  as  forty  times.  Different  colloids  differ  greatly  in 
their  affinity  for  water,  and  in  the  way  in  which  this  affinity  is  mod- 
ified by  electrolytes,  and  change  in  a  colloid  may  greatly  alter  its  ca- 
pacity for  swelling;  thus,  ;8-gelatin,  which  can  be  formed  from  ordi- 
nary gelatin  by  the  action  of  proteolytic  enzymes,  has  greater  capacity 
for  swelling  than  the  original  gelatin.  Gies  especially  lays  stress  on 
this  factor,  that  is,  the  alterations  of  the  hydrophilic  tendencies  of  the 
tissue  colk)ids  by  enzymes. ^^ 

On  the  basis  of  the  facts  briefly  summarized  above,  the  proportion 
of  water  present  in  any  cell  or  in  any  fluid  of  the  body  which  contains 
colloids,  is  assumed  to  be  determined  by  certain  factors,  namely  (1) 
the  cliaracter  of  the  colloids  themselves;  (2)  the  proportion  and  na- 
ture of  acids  or  alkalies  present  in  the  fluids  in  and  about  the  colloids ; 
(3)  the  proportion  and  nature  of  the  salts.  All  these  factors  are 
changeable,  and  tlierefore  the  amount  of  water  present  in  the  cell  or 
fluid  varies  accordingly.  Thus,  if  a  cell  through  its  metabolism  de- 
velops from  such  a  non-electrolyte  as  sugar  (which  has  no  consider- 
able effect  on  the  water  content  of  the  protoplasm),  an  organic  acid, 

i3aAmer.  Jour.  Physiol.,  in07    (10),  .-^eO:    1008    (22),  01. 

1*  See  Fischer's  INlonopraph,  "Oedema  and  Nepliritis."'  New  York.  1015;  also 
numerous  articles  in  tlie  Zeit.  f.  Chem.  u.  Ind.  d.  Kolloide.  An  especially  thor- 
ou^'h  discussion  of  this  theory  is  contained  in  the  biochemical  l^villctin.  Vol.  T., 
pivinj^  a  bihliofijraphy  of  Fischer's  work,  together  with  articles  on  Gies'  observa- 
tions on  the  modification  of  the  hydrophilic  tendency  of  proteins  by  enzyme 
action. 

1' A  definite  and  clear-cut  example  of  tlu^  swelling'  of  a  tissue  under  the  in- 
fluence of  acid  of  metabolic  orifjin  is  shown  in  the  muscle  cell  in  Zenker's  waxy 
degeneration    (Wells,  Jour.  Exper.  Med.,   1000    (11),  1). 


FORMATION  OF  JAMI'JI  337 

such  as  lactic  acid,  which  has  a  large  effect  in  increasing  the  affinity 
of  tile  colloids  for  water,  the  cell  will,  presumably,  take  on  more  water, 
perhaps  to  a  degree  to  cause  intracellular  edema.  The  acids  diffusing 
from  the  cell  into  the  intercellular  spaces  or  into  the  lymph  will 
cause  equally  well  an  increased  affinity  for  water  in  the  colloids  here 
present,  leatling  to  intercellular  edema.  Conversely,  neutralization 
of  acids  present  in  a  colloidal  solution,  by  alkaline  salts  brought  by 
the  blood,  will  decrease  the  affinity  of  the  colloids  for  water  which 
will  escape  from  the  colloids  as  they  shrink. 

This  theory,  which  introduces  a  hitherto  unappreciated  factor  into 
the  considerations  of  lymph  formation  and  edema,  is  of  the  utmost 
importance.  It  practically  eliminates  osmotic  pressure,  also  the  cell 
membranes  so  essential  for  the  efficiency  of  this  force,  and  in  view  of 
the  difficulties  that  have  arisen  in  trying  to  fit  the  cell  membrane 
hypothesis  and  osmotic  pressure  to  many  facts  of  normal  and  patho- 
logical biology,  an  alternative  hypothesis  is  welcome.  As  pointed  out 
above,  the  forces  involved  in  the  swelling  of  colloids  are  so  large  as  to 
be  of  great  significance,  and  the  amounts  of  electrolytes  necessary 
to  cause  considerable  variations  in  colloidal  swelling  are  not  more  than 
can  be  present  under  normal  and  pathological  conditions;  conse- 
quently the  possible  influence  of  colloidal  swelling  must  be  taken 
into  account  in  all  consideration  of  pathological  processes.  Whether 
or  not  it  is  capable  of  as  universal  application  as  Fischer  maintains, 
remains  to  be  demonstrated,  and  there  are,  indeed,  some  facts  that 
do  not  seem  to  be  in  harmony  with  this  theory. 

Summary. — We  see  from  the  above  discussion  that  numerous  the- 
ories have  been  advanced  to  explain  the  normal  formation  of  lymph, 
and  as  their  basis  exist  several  different  possible  factors.  Filtration, 
active  secretion  by  the  capillary  endothelium,  attraction  by  the  tissue- 
cells,  osmosis  in  response  to  formation  of  crystalloids  outside  the  ves- 
sels, and  changes  in  the  affinity  of  colloids  for  water;  all  have  been 
shown  to  be  possible  causes  of  lymph  formation.  It  is  highly  prob- 
able that  in  a  certain  way  all  are  involved,  particularly  if  we  accept 
the  view  of  the  physical  school  that  "secretion"  and  "attraction" 
by  the  cells  are  merely  the  outcome  of  physical  forces ;  the  causes  of 
lymph  formation  then  reduce  themselves  to  absorption,  filtration  and 
diffusion.  There  has  been,  until  recenth-,  no  question  but  that  lymph 
does  escape  from  the  vessels  through  simple  filtration,  for  the  pressure 
inside  the  capillaries  is  presumably  greater  than  outside,  the  capil- 
lary walls  are  not  water-tight,  and  the}^  are  not  impermeable  to  the 
substances   dissolved    in    the    plasma.^'     Likewise    osmotic    exchanges 

17  Hill  ("Recent  Advances  in  Physiolo^iy  and  Biochemistry,"  1000,  p.  GIS)  dis- 
putes tlie  possibility  of  such  a  thing  as  filtration  pressure,  on  the  <rroiuid  that 
the  structures  within  the  capsule  of  an  orfian  must  all  be  under  the  influence 
of  the  blood  pressure  alike;  but  with  the  presence  of  an  outlet  for  tlie  fluid,  as 
in  glands  with  ducts,  filtration  pressure  surely  can  applv. 
22 


338  EDEMA 

surely  go  ou  between  the  vessels  aud  the  tissue-cells,  and  the  condi- 
tions which  determine  the  water  content  of  our  colloid  solutions 
constantly  vary.  The  question  that  remains  is,  do  these  two  factors 
account  for  all  of  the  lymph  formation,  and  are  they  sufficient  by 
themselves  to  explain  the  physiological  regulation  and  the  pathological 
variations  in  the  Ij'mph  flow?  They  are  purely  physical  or  mechan- 
ical causes,  and  the  "vitalist"  school  will  claim  that  they  are  inade-  ^ 
quate  and  that  "vital  activities"  of  the  cells  play  the  deciding  role. 
But  at  present  the  evidence  that  is  being  accumulated  seems  to  point 
more  and  more  strongly  to  the  conclusion  that  these  ' '  vital  activities ' ' 
are  but  the  result  of  simple  well-known  physical  forces  acting  under 
very  complex  conditions — complex  because  of  the  large  number  of 
different  chemical  compounds  occurring  together,  and  the  varying  in- 
fluence of  circulation,  food  supplies,  cell  structure,  etc. 

ABSORPTION  OF  LYMPH 

By  no  means  all  the  fluid  that  escapes  from  the  vessels,  nor  all  the 
products  of  cell  metabolism,  are  carried,  away  in  the  lymph — a  con- 
siderable and  perhaps  the  greater  part  of  them  is  absorbed  back 
into  the  capillaries  directly.  A  classical  proof  of  this  is  the  experi- 
ment of  Magendie,  who  observed  that  if  poisons  were  injected  into  the 
leg  of  an  animal,  which  had  been  separated  from  the  body  entirely 
except  for  the  blood-vessels,  that  poisoning  developed  in  the  usual 
manner.  In  such  experiments  the  lymph-vessels  are  severed  and  prob- 
ably largely  occluded ;  hence  it  does  not  solve  the  question  as  to 
wiiether  substances  are  absorbed  by  the  blood-vessels  under  normal 
conditions.  Orlow  found,  however,  that  during  absorption  of  fluid 
from  the  peritoneal  cavity  there  is  no  perceptible  increase  in  the 
lymph  flow  from  the  thoracic  duct.  Addition  of  sodium  fluoride,  a 
protoplasmic  poison,  was  found  to  interfere  with  this  absorption,  for 
which  and  other  reasons  Heidenhain  and  Orlow  considered  that  the 
absorption  depended  upon  the  "vital  activity"  of  the  cells.  ^Fore 
nearly  reproducing  normal  conditions  were  the  experiments  of  Star- 
ling and  Tubby,  who  found  that -methylene-blue  or  indigo-carmine  in- 
jected into  the  pleura  or  peritoneum  appeared  in  th(^  urine  long  be- 
fore it  colored  the  lymph  in  the  thoracic  duct.'**  Adler  and  Meltzer 
found  evidence,  however,  that  not  all  the  absorption  is  accomplished 
by  the  blood-vessels,  for  obstruction  of  the  thoracic  duct  retards  ab- 
sorption. That  the  absorption  is  not  dependent  solely  upon  the  cir- 
culation and  blood  pressure  is  shown  by  the  fact  tiuit  absorption 
from  the  peritoneal  cavity  occurs  in  dead  bodies  (Hamburger,  Adler 
and  Meltzer). 

The  nature  of  the  mechanism  l)y  wliidi  Huids  arc  taken  into  tlie 
blood-vessels  is  still  unknown.  We  can  easily  understand  tlie  en- 
trance of  injected  poisons  and  eoloring-mattei-s  from  the  tissues  into- 

isStHJ  Mendel,  Amer.  Jour.  Physiol.,  1899   (2),  342. 


THE  CAUSES  OF  EDEMA  339 

the  blood,  because  they  are  more  concentrated  at  the  point  of  injection 
than  in  the  bh)()d,  hence  they  may  diffuse  directly  through  the  capil- 
lary wall.  Likewise  we  can  understand  the  diffusion  of  water  from 
a  hypotonic  solution  into  the  blood,  but  how  a  solution  of  the  same 
concentration  as  that  of  the  blood  can  enter  the  blood  is  difficult  to  ex- 
plain. Cohnstein  and  also  Starling  attribute  this  absorption  to  the 
proteins  of  the  blood  in  the  following  manner:  After  a  fluid  is  in- 
jected into  the  tissues  or  serous  cavities  there  occurs  a  diffusion  ex- 
change between  this  fluid  and  the  blood,  until  the  concentration  of  the 
crystalloids  in  each  is  equal;  but  the  proteins  of  the  blood  cannot 
diffuse,  and  as  they  exert  a  positive  although  very  slight  osmotic  pres- 
sure, this  difference  in  osmotic  pressure  in  favor  of  the  blood  causes 
diffusion  of  the  extravascular  fluid  into  the  blood.  Roth  has  also  ap- 
plied this  idea  in  a  rather  complicated  manner  to  the  absorption  oc- 
curring in  metabolic  processes  (see  Meltzer),  but  it  must  be  admitted 
that  it  is  an  unsatisfactory-  solution  of  the  problem.  Fischer  would 
ascribe  the  passage  of  fluid  to  the  relative  affinity  of  the  colloids  of  the 
blood  and  of  the  tissues  for  the  fluid,  and  this  would  be  towards  the 
blood  whenever  the  blood  colloids  had,  from  whatever  possible  cause, 
a  greater  affinity  for  the  fluid  than  the  tissue  colloids. 

Passage  of  the  fluid  from  the  tissues  into  the  lymph  stream  was  very 
easy  to  understand  in  the  light  of  the  older  conception  of  the  lym- 
phatic circulation,  namely,  that  the  lyinph-vessels  were  merely  con- 
tinuations of  the  interstitial  spaces ;  we  could  then  assume  that  as 
soon  as  the  fluid  left  the  blood-vessels  it  was  practically  within  the 
lymphatic  system,  and  was  crowded  along  the  Ij-mphatic  channels  by 
the  vis  a  tergo,  aided  by  the  valves  of  the  lymph-vessels  and  the  intra- 
thoracic vacuum.  But  it  now  seems,  particularly  through  the  studies 
of  MacCallum.^'^  that  the  lymphatic  vessels  form  a  closed  system,  not 
in  communication  with  the  interstitial  spaces.  This  being  the  case, 
we  have  to  explain  the  passage  of  the  lymph  through  the  walls  of 
the  lymphatic  vessels,  and  this  is  a  problem  which  is  not  by  any 
means  a  simple  one,  and  which  has  yet  to  be  investigated.  It  is  sig- 
nificant that  the  thoracic  lymph  has  a  higher  osmotic  pressure  than 
the  blood  of  the  same  animal  (Luckhardt),-"  so  that  the  lymph  which 
enters  the  duct  must  do  so  against  the  osmotic  pressure. 

THE  CAUSES  OF  EIEMA 

With  the  facts  and  hypotheses  mentioned  in  the  preceding  para- 
graphs in  mind,  we  may  consider  their  bearing  on  the  production  of 
abnormally  large  accumulations  of  lymph  in  the  tissues,  that  is,  edema. 
We  can  imagine  any  one  of  the  following  factors  as  causing  or  helping 
to  cause  such  a  pathological  accumulation: 

19  Johns  Hopkins  Hosp.  Bull.,  1903    (14),  105. 
20Anier.  Jour.  Physiol.,  1910   (25),  345. 


340  EDEMA 

.    1,  Obstruction  to  outflow  through  the  lymph-vessels. 

2.  Increased  blood  pressure. 

3.  Decreased  extravascular  pressure. 

4.  Increased  permeability  of  the  capillary  walls. 

5.  Increased  filterability  of  the  blood  plasma. 

6.  Osmotic  pressure  changes — either  an  extravascular  increase  or 

an  intravascular  decrease. 

7.  Changes  in  the  affinity  of  the  colloids  for  water. 

These  may  be  taken  up  one  by  one,  and  considered  in  relation  to 
their  bearing  upon  the  general  problem  of  edema. 

1.  Obstruction  to  Outflow  through  the  Lymph=vessels. — ^Be- 
cause of  the  very  abundant  anastomosis  of  the  lymphatic  vessels  it  is 
extremely  difficult  or  impossible  to  cause  any  appreciable  obstruction 
to  the  lymphatic  circulation  by  ligation  of  lymphatic  trunks  in  the 
limbs  or  organs  of  the  body,  and  in  pathological  conditions  this  possi- 
ble cause  of  edema  is  seldom  actually  observed.  The  chief  instance 
of  edema  from  lymphatic  obstruction  is  observed  after  occlusion  of  the 
thoracic  duct  by  tumors,  tuberculous  processes,  animal  parasites,  or 
thrombosis ;  such  occlusion  is  usually  followed  by  rupture  of  the  duct 
or  its  tributaries,  with  the  production  of  chylous  ascites  or  chylothorax, 
and  chyluria.  Filaria  or  their  ova  may  occupy  so  many  of  the  lymph- 
atic channels  of  an  extremity  (leg)  or  part  (scrotum)  that  the  an- 
astomotic channels  are  thoroughly  blocked,  with  a  resulting  local 
edema  that  in  course  of  time  is  followed  by  the  production  of  in- 
flammatory connective  tissue  and  elephantiasis.-^  Chronic  lymph- 
angitis or  plugging  of  the  lymph  vessels  by  cancer  cells  may  also  result 
in  lymphatic  obstruction  to  such  an  extent  that  chronic  edema  results. 
It  would  seem,  from  Opie's  experiments,-'-  that  the  acute  edemas  may 
at  times  depend  upon  lymphatic  obstruction,  for  he  found  that  experi- 
mental edema  of  the  liver,  produced  by  cantharidin,  seems  to  be 
determined  by  inflammatory  processes  which  occlude  the  sinuses  of 
the  lymph  glands  through  which  the  hepatic  lymph  passes. 

Another  way  in  which  edema  may  be  caused  or  influenced  by  lympli- 
atic  obstruction  is  generally  overlooked,  but  it  is  possibly  of  great 
importance;  namely,  from  pressure  upon  the  lymph  channels  by 
dilated  vessels  in  hyperemia,  or  by  cellular  exudates  and  swollen 
tissues  in  inflamnuition.  We  see  evidence  of  this  in  the  rapid  absorp- 
tion of  exudates  that  frequently  follows  the  removal  of  but  a  part  of 
the  fluid  in  a  chest  cavity ;  apparently  the  decrease  in  pressure  frees 
the  paths  of  absorption  and  permits  them  to  take  up  the  remaining 
fluid.  In  inflammatory  edema  tlie  lym])hatic  obstruction  is  probably 
not  great,  for  Lassar  found  tliat  the  amount  of  lym|)h  escaping  from 
an  edematous  extremity  is  inncli   gi-eatei-  than    Ironi  a   normal   one; 

siManson,  Allhutt's  Svstoin,  1S97    (ii),  1082. 
i:a  Jour.  Exper.  Med.,  id]2   (16),  8.31. 


THE  CAUSES^  OF  EDEMA  341 

but  in  the  case  of  strangulated  hernias  or  other  conditions  in  wliich 
edema  results  from  circular  constriction,  obstruction  of  the  lymphatic 
vessels  may  be  a  factor  of  no  mean  importance.  In  general  stasis  the 
increased  pressure  in  the  veins  of  the  neck  may  interfere  with  the 
passage  of  the  fluid  out  of  the  thoracic  duct  into  the  blood. 

There  is  no  difficulty  in  understanding  edema  from  the  above  causes 
— it  is  simply  a  passive  congestion  of  the  lymphatic  circulation,  and  no 
chemical  factors  are  involved.  The  nature  of  the  fluid  found  in  such 
forms  of  edema  will  be  discussed  later. 

2.  Increased  Blood  Pressure. — This  takes  us  back  to  the  filtra- 
tion theoiy  of  lymph  formation,  and  as  it  is  generally  conceded  that 
more  or  less  fluid  escapes  from  the  vessels  by  this  mechanical  process, 
the  questions  to  be  decided  are :  Can  and  does  increased  blood  pres- 
sure, alone  and  without  other  aiding  factors,  cause  edema?  If  not, 
does  it  play  an  auxiliary  part  in  producing  edema,  and  how  important 
a  part  may  this  be?  Many  experiments  have  been  performed  with  the 
object  of  answering  these  questions,  with  more  or  less  conflicting  re- 
sults. Cohnheim  demonstrated  that  vasodilation  (active  hyperemia) 
alone  will  never  bring  on  an  edema ;  and  many  observers  state  that 
ligation  of  the  femoral  or  other  large  veins  will  not  cause  edema  in 
animals.  However,  when  the  vein  is  occluded,  and  the  arteries  are 
dilated  by  cutting  their  vasoconstrictor  nerves,  then  edema  may  result 
(Ran\aer,  Cohnheim)  ;  but  whenever  venous  outflow  is  impeded,  we 
have  other  factors  than  simply  increased  pressure  to  consider,  for  the 
nourishment  of  the  parts  is  decidedly  impaired,  and,  as  we  shall  see 
later,  this,  may  be  of  much  greater  importance  than  is  the  associated 
rise  in  blood  pressure.  To  produce  edema  in  the  lungs  by  mechanical 
forces  it  is  necessary  to  ligate  the  aorta  and  its  branche^s,  or  the  pul- 
monary veins  (Welch).  As  such  high  pressures  do  not  occur  in  any 
pathological  conditions,  it  is  safe  to  assume  that  increased  pressure 
alone  is  not  capable  of  causing  by  itself  the  pulmonary  edema  so  fre- 
quently' observed  clinically.  AVelch,'^  however,  has  supported  the 
hypothesis  that  ..a  disproportion  between  the  working  power  of  the 
left  ventricle  and  of  the  right'  ventricle  may  lead  to  pulmonary  edema 
through  pulmonary  hyperemia.  In  the  edema  of-passive  congestion, 
increased  blood  pr-eJ*5ure'^woul{i  seem  to  be  an  impoj'tant  factor,  and 
there  is  ^o  (^ohbt  tha,t  with  ^n  increased  pressiia;^  of  the  degree' 
observed  in  such  conditions  s(5me  increase- in  the\ljniiph  floVv  would 
result ;  but  from  the  evidence  at  hand  it  is  •improbarrle  -that  the  amount^- 
of  lymph  so  secreted  woulft.gner  be  more  than  the  lyniph-vessels  could^ 
carry  away.  Even  the  added  obstruction  to  lymphatic  flow  due  to 
pressure  upon  the  lymph  capillaries  by  congested  blood-vessels,  and 
the  resistance  to  the  lymph  escaping  from  the  thoracic  duct  offered 
by  the  increased  pressure  in  the  subclavian  vein,  would  not  satisfac- 
torily account  for  the  edema  of  cardiac  incompetence.     Not  to  go  into 

zsVirchow's  Arch.,  1878  (72),  375;  see  also  Meltzer  {loc.  cit.) . 


342  EDEMA 

details  here,  it  ma}'  be  stated  that  the  impression  is  growing  that  un- 
complicated rise  in  blood  pressure  is  not  sufficient  by  itself  to  pro- 
duce edema.  Some  of  the  reasons  for  belittling  this  factor  will  be 
brought  out  in  the  subsequent  discussion. 

3.  Decreased  Extravascular  Pressure. — This  factor  is  particu- 
larlj'  prominent  in  the  so-called  "'edema  ex  vacuo,"  which  occurs  after 
the  absorption  of  an  area  of  tissue  which  is  so  located  that  the  sur- 
rounding tissues  cannot  contract  or  fall  in  to  fill  the  gap,  e.  g.,  brain 
softening,  serous  atrophy  of  fat.  A  still  better  example,  however, 
is  the  edema  that  follows  local  decrease  in  atmospheric  pressure  in 
"cupping."  In  these  instances  the  edema  depends  partly  upon  in- 
creased transudation,  and  partly  on  the  retention  of  the  fluid  in  the 
tissues,  because  it  cannot  well  leave  them  against  the  atmospheric 
pressure.  The  idea  advanced  by  Landerer  that  decreased  elasticitj^ 
of  the  tissues  was  a  possible  cause  of  edema  has  been  attacked  by  Bon- 
niger,-*  who  found  but  little  alteration  in  the  elasticity  of  tissues  the 
seat  of  edema.  During  the  early  stages  of  edema,  however,  the  elas- 
ticity of  the  skin  may  be  measurably  decreased,-^  even  when  no  edema 
is  demonstrable  by  palpation;  but  this  is  not  evidence  that  any  loss 
of  elasticity  occupies  a  causative  relation  to  the  edema.  The  tissues 
can  take  up  water  until  as  much  as  six  kilos  has  been  added  to  the 
Aveight  of  the  entire  body  before  any  edema  can  be  detected  by  pal- 
pation (Widal).  Edema  ex  vacuo  is  again  an  illustration  of  edema 
due  to  purely  mechanical  causes,  but  it  is  of  little  practical  impor- 
tance. 

4.  Increased  Permeability  of  the  Capillary  Walls. — The  im- 
portance of  this  factor  in  the  production  of  edema  was  first  demon- 
strated by  Cohnheim  and  Lichtheim.  who  found  that  the  production 
of  an  enormous  increase  in  the  amount  of  fluid  in  the  blood  (hydremic 
plethora)  by  injecting  large  quantities  of  salt  solution,  caused  an 
edema  of  the  viscera  and  serous  cavities,  ])ut  not  any  subcutaneous 
edema  until  the  skin  had  been  irritated  by  some  means,  such  as  hot 
water,  iodin,  etc-.  By  this  irritation  the  ca])illary  walls  are  injured, 
and  an  excessive  escape  of  the  blood  fluids  follows,  ^lagnus  also 
showed  that  poisoning  with  arsenic,  which  injured  the  vessels,  favored 
the  experimental  production  of  edema  by  transfusion.  Starling,  as 
noted  before,  observed  that  the  permeability  of  the  ca])illaries  varies 
normally  in  ditferent  organs  and  tissues,  which  determines  quantita- 
tive and  qualitative  ditferences  in  the  lymph  normally  flowing  from 
various  vascular  areas.  Heidenhain's.  "lymphagogues  of  the  fii*st 
class,"  which  are  all  poisonous  substances,  probably  act  by  increasing 
the  permea])ility  of  the  cainllaries,  and  in  this  way  they  produce 
local  iirticaria,  which  is  often  observed  as  a  result  of  iioisoning  by 
these  same  lymphagogues,  e.  f/.,  shellfish   and   strawhci-iy   poisoning. 

aiZoit.  oxp.  Path.  u.  Thor.,  1005    (1).  lU.'^. 
ssSchade,  Zeit.  exp.  Patli.,   1012    (11),  360. 


THE  CAUSES  OF  EDEMA  343 

Just  wliat  chaii're.s  are  produced  in  tlie  capillary  walls  that  render 
them  more  i)ermeal)le  we  do  not  know.  Possibly  in  some  instances  it 
is  a  partial  solution  of  the  intei-eellular  cement  substances,  possibly 
an  enlargement  of  the  stomata  through  loss  of  tonicity  of  the  endo- 
thelium (^Meltzer),  sometimes  it  may  be  actual  death  of  the  endothelial 
cells,  or,  as  Heidenhain  and  Cohnheira  thought,  it  may  be  a  stinuila- 
tion  of  the  endothelial  cells  to  increased  secretory  activity.  Fischer 
believes  that  a  change  in  the  hydrophilic  tendency  of  the  colloids, 
induced  especially  by  acids  formed  in  asphyxiated  conditions 
of  the  colls,  alter  their  structure  and  with  that  their  permeabil- 
ity. 

Under  pathological  conditions  increased  permeability  of  the  capil- 
lary walls  is  probably  one  of  the  chief  factors  in  the  production  of 
certain  forms  of  edema.  We  see  evidence  of  it  particularly  in  inflam- 
matory edema,  with  its  protein-rich  exudate.  It  cannot  be  doubted 
that  in  such  conditions  actual  physical  alterations  take  place  in  the 
capillaries,  when  we  see  that  the  slightly  diffusible  proteiijs  escape 
from  the  vessels  in  the  same  proportions  as  th&y  exist  in  the  plasma; 
there  can  be  here  no  question  of  heightened  cell  activity  or  increase  in 
osmotic  pressure,  especially  not  Avhen  Ave  note  the  indistinguishable 
transition  of  such  an  inflammatory  exudate  into  one  containing  leu- 
cocytes and  red  corpuscles,  which  must  pass  through  openings  of 
some  kind  in  the  vessels.  Edema  due  to  inflammation  and  poisoning 
certainly  depends  to  a  large  degree  upon  alterations  in  the  vessel- 
walls.  The  question  remaining  is,  do  edemas  that  are  not  asso- 
ciated with  distinct  inflammatory  or  toxic  influences  depend  also  upon 
the  vascular  permeability? — does  increased  permeability  ever  lead  to 
the  formation  of  protein-poor  transudates?  Cohnheim  was  inclined 
to  attribute  nearly  all  edema  to  this  cause,  for  in  passive  congestion, 
or  nephritis,  or  any  of  the  common  causes  of  edema,  it  is  easy  to  find 
reason  for  the  belief  that  poisons  may  be  present  in  the  blood;  and 
as  there  w:as  good  evidence  that  the  blood  pressure  alone  could  not 
account  foKthe  edema,  it  was  natural  to  Ascribe  all  these  fornYs  of 
edema  to  the  action  of  toxic  substances  upon  the  capillary  walls,  lead- 
ing to  increased  permeability;  or,  what  might  amount  to  the  same 
thing,  increased  secretory  activity  of  the  endothelium,  as  understood 
by  Heidenhain.  It  is  impossible  at  this  time  to  eliminate  as  yihn-  ,  ; 
existent  this  secretory-activity  doctrine,  but,  as  we  hope  to  show  later-, 
there  exist  other  factors  in  all  these  non-inflammatmy  edtniQ^f^tlTat^"* 
are  sufficient  to  account  for  the  edema  without^ur  ha^*ig  r^jMirseto 
this  hypothesis.  For  the  present,  therefare.  wt^-^jiay  con>n#ler  altered 
capillary  permeability  as  an  essential  factor  in  edema^^  characterized 
by  protein-rich  fluids  (exudates),  and  iglc'ffe^that'.tlie  influence  of  al- 
tered permeability  in  the  producti^*  of  protein-poor  fluids  (trans- 
udates) is  not  proved,  and  is  perhaps  not  of  importance,  although 
the   evidence  of  recent  studies  on-  experimental  nephritis  seems  to 


344  EDEMA 

point  more  and  more  to  the  importance  of  vascular  chang:es  in  acute 
nephritis,  at  least.-" 

5.  Increased  Filterability  of  the  Blood  Plasma. — This  takes 
us  back  to  Richard  Bright 's  conception  of  renal  dropsy.  He  im- 
agined that  through  the  great  loss  of  albumin  in  the  urine  the  blood 
became  so  thinned  and  watery  that  it  could  filter  through  the  vessel- 
walls,  while  normal  plasma,  he  thought,  was  too  thick  and  viscid  to  do 
so.  The  same  idea  was  applied  to  the  edemas  of  cachexia  in  cancer, 
etc.,  chlorosis,  and  all  forms  of  edema  associated  with  a  decrease  in 
the  corpuscular  or  protein  elements  of  the  blood.  With  our  present 
knowledge  of  diffusion  of  crystalloids  and  colloids  we  can  readily  ap- 
preciate that  a  decrease  in  the  blood  colloids,  such  as  might  occur  in 
these  diseases,  could  not  modify  the  passage  of  fluids  through  the 
capillary  walls  to  any  considerable  degree.  Stewart  and  Bartels  con- 
sidered that  in  renal  dropsy  the  increased  filterability  of  the  plasma 
was  not  due  so  much  to  the  loss  in  albumin  as  to  retention  of  water, 
which  caused  an  hydremic  plethora.  But  this  factor  was  soon  elimi- 
nated, for  it  was  found  that  complete  anuria,  produced  by  ligating 
both  ureters,  does  not  cause  edema ;  and  also  that  to  produce  an  edema 
by  increasing  the  water  of  the  blood  it  was  necessary  to  increase  it 
many  times  as  much  as  it  can  ever  be  increased  by  disease.  Simpl}^ 
increasing  the  proportion  of  water  by  removing  part  of  the  blood  and 
injecting  a  corresponding  amount  of  salt  solution  did  not  cause  edema 
iCohnheim  and  Lichtheim).  We  may,  therefore,  look  upon  the  hy- 
pothesis of  increased  filterability  of  the  blood  as  chiefly  of  historic 
interest,  and  not  an  important  factor  in  the  causation  of  edema.  In 
the  presence  of  other  factors  for  the  production  of  edema,  however,  the 
amount  of  fluid  in  the  vessels  is  important;  thus  Pearce -'  found  that 
in  experimental  uranium  nephritis  hydremia  exerted  a  marked  in- 
fluence on  the  production  of  edema. 

6.  Disparity  of  Osmotic  Pressure  in  Favor  of  the  Tissues  and 
Lymph  over  the  Blood. — On  a  preceding  page  we  have  already 
considered  the  means  by  which  changes  in  osmotic  pressure  in  the  tis- 
sues are  brought  about,  and  how  they  may  lead  to  an  accumulation 
of  fluid.  The  importance  of  osmotic  pressure  in  causing  pathological 
edema  was  suggested  by  J.  Loeb  -^  in  his  studies  on  the  physiological 
action  of  ions.  He  stated  that  edema  occurred  when  the  osmotic 
pressure  was  higher  in  the  tissues  than  it  was  in  the  blood  and  lympli, 
and  the  cause  was  to  be  sought  in  conditions  that  lowered  the  osmotic 
pressure  of  the  blood  and  lympli  or  raised  that  of  the  tissues.  This 
condition  he  found  in  the  accumulation  of  metabolic  products : — in 
the  case  of  muscle,  totanization  of  a  fi-og's  muscle  for  ten  minutes 
raised  the  osmotic  pressure  over  one  atmosphere ;  separating  a  muscle 

26  See  Schmid  and  Sdilavcr,  Dent.  Arch.  klin.  Med.,  1011    (104).  44. 

27  Arch.  Int.  Med.,  1908  "(3),  422. 
28Pfluger's  Arch.,  1898   (71),  457. 


THE  CAUSES  OF  EDEMA  345 

from  its  l)l()()(l-supi)ly  led  to  sueli  an  increase  in  osmotic  pressure  tliat  it 
took  uj)  water  from  a  4.!)  per  cent.  NaCl  solution,  wliich  lias  a  j)res- 
sure  of  over  thirty  atmospheres.  AVhen  we  consider  that  in  his  studies 
on  lung  edema  Welch  was  able  by  ligation  of  the  aorta  to  raise  the 
blood  pressure  less  than  ^^o  atmosphere,  we  begin  to  appreciate  how 
much  more  powerful  are  the  physico-chemical  forces  that  are  at  work 
in  the  body  than  is  the  blood  pressure,  even  of  the  aorta  itself. 
^  Loeb  found  that  whenever  oxidation  is  impaired  in  a  tissue  its 
osmotic  pressure  rises,  which  he  ascribed  to  the  accumulation  of  in- 
completely oxidized  metabolic  products,  particularly  acids,  and  as  a 
result  the  muscle  takes  up  water  and  becomes  edematous.  On  this 
basis  we  might  explain  the  edema  of  venous  stagnation  as  due  to  ac- 
cumulation of  products  of  metabolism,  partly  because  of  impaired 
oxidation,  partly,  perhaps,  because  of  their  slow  removal  in  the  blood 
on  account  of  the  circulatory  disturbance.  The  so-called  ''neurotic" 
edemas  may  possibly  be  explained  by  local  increase  in  metabolic  ac- 
tivity brought  about  by  nervous  stimuli,  which  causes  increased  forma- 
tion of  substances  raising  osmotic  pressure  in  the  stimulated  tissues. 
In  renal  edema  the  retention  of  water  also  seems  to  depend  rather  on 
osmotic  pressure  than  on  circulatory  disturbances  or  alterations  in  the 
vessel- walls,  for  it  has  been  shown  that  retention  of  chlorides,  which 
the  diseased  kidneys  do  not  eliminate  normally,  is  an  important  cause 
of  the  dropsy  in  some  cases.  The  chlorides  accumulating  in  the  tissues 
lead  to  an  increased  osmotic  pressure,  which  causes  the  abstraction  of 
water  from  the  blood  and  its  retention  in  the  tissues.  (The  details  of 
this  subject  will  be  considered  later.)  Convereely,  Meltzer  and  Salant 
found  that  salt  solution  is  absorbed  from  the  peritoneal  cavity  more 
rapidly  in  nephrectomized  rabbits  than  in  normal  rabbits,  because 
metabolic  products  accumulate  in  the  blood  and  raise  its  osmotic  pres- 
sure above  normal ;  and  it  was  observed  by  Fleisher  and  L.  Loeb  ^^ 
that  the  rate  of  absorption  of  fluid  from  the  peritoneal  cavity  is  in- 
creased when  the  osmotic  pressure  of  the  blood  is  raised. 

There  are  some  difficulties,  however,  in  applying  the  influence  of 
osmotic  pressure  as  an  explanation  of  all  edemas.  For  example,  in 
edema  of  the  lungs,  as  Meltzer  points  out,  what  is  the  force  that  drives 
the  fluid  into  the  empty  air-cells?  Equally  difficult  to  explain  as  the 
result  of  osmotic  disturbance  is  the  distribution  of  fluid  that  is  seen 
in  cardiac  dropsy.  The  fluid  does  not  accumulate  in  the  tissues  where 
metabolism  is  greatest,  or  where  the  most  oxygen  is  used ;  but  rather 
in  the  inactive  subcutaneous  tissues  and  in  the  serous  cavities.  Possi- 
bly the  original  transudation  does  occur  in  the  muscles  and  solid 
viscera,  and  the  fluid  is  then  mechanically  forced  out  of  them  into 
the  surrounding  tissue-spaces,  later  settling  according  to  the  laws  of 
gravity  or  according  to  the  distensibility  of  the  tissues.  It  is  im- 
portant to  take  into  consideration  the  fact  that  demonstrable  edema 
29  Jour.  Exper.  Med.,  1910  (12),  510. 


346  EDEMA 

does  not  manifest  itself  until  a  very  large  quantity  of  fluid  has  been 
retained  by  the  body — as  much  as  six  kilos,  according  to  AVidal. 

Increased  Hydration  Capacity  of  the  Tissue  Colloids. — According  to 
Fischer's  tlieory  this  factor  is  of  greater  importance  than  any  of  the 
preceding,  and  of  chief  importance  in  increasing  the  amount  of  water 
present  in  the  tissues  are  organic  acids  formed  during  metabolism. 
For  example,  the  great  power  of  asphyxiated  muscle  to  take  up  water 
from  a  strong  salt  solution,  which  J.  Loeb  ascribed  to  the  osmotic, 
pressure  of  the  acids  formed  in  asphyxia,  is  attributed  by  Fischer 
to  the  influence  of  tliese  acids  upon  the  capacity  of  the  colloids  for 
water,  and  this  explanation  seems  to  be  in  better  agreement  with  the 
facts,  especially  since  Overton  has  shown  that  even  if  all  tlie  proteins, 
earboliydrates  and  fats  in  a  muscle  were  split  into  the  greatest  possi- 
ble number  of  simple  molecules  and  ions,  the  resulting  osmotic  pres- 
sure would  not  be  sufficient  to  account  for  the  amount  of  water  taken 
up.  Furthermore,  when  cells  with  demonstrable  semi-permeability 
die,  they  at  once  lose  their  semi-permeability,  and  in  consecjuence  their 
osmotic  pressure  falls — but  dead  cells  and  tissues  often  exhibit  great 
power  of  taking  up  water  and  becoming  edematous.''"  It  is  an  in- 
disputable fact  that  edema  is  especially  associated  Math  conditions  of 
asphyxiation,  and  the  attempt  to  explain  this  by  the  increased  osmotic 
pressure  of  the  products  of  incomplete  oxidation  seem  to  harmonize 
with  the  facts  far  less  successfully  than  the  application  of  the  prin- 
ciple of  colloidal  swelling.  A  common  error  of  the  critics  of  this 
theory  is  that  of  assuming  that  free  acid  must  be  present  to  cause 
swelling.  This  is  not  at  all  true.  An  amount  of  acid  far  less  than 
enough  to  saturate  the  acid-binding  property  of  a  protein  or  to  be 
detected  by  indicators  will  greatly  increase  the  amount  of  water  which 
this  protein  will  combine.  Presumably  the  colloidal  carbohydrates  and 
lipoids  may  also  play  a  part  in  the  water  absorption  of  tissues. 

Fischer's  theory  of  edema,  in  his  own  words,  is  this:  "A  state  of 
edema  is  induced  whenever,  in  the  presence  of  an  adequate  supply 
of  water,  the  affinity  of  the  colloids  of  the  tissues  for  water  is  in- 
creased above  that  which  we  are  pleased  to  call  normal.  The  ac- 
cumulation of  acids  within  the  tissues  brought  about  either  through 
their  abnormal  production,  or  through  the  inadequate  removal  of  such 
as  some  consider  normally  produced  in  the  tissues,  is  chiefly  responsi- 
ble for  this  increase  in  the  affinity  of  the  colloids  for  water,  though 
the  possibility  of  explaining  at  least  some  of  the  increased  affinity  for 
water  tlirough  the  production  or  accunnilation  of  substances  which  af- 

^0  The  secreted  fluid  of  postmortem  tlioraeie  lymiili  How  dilVers  from  normal 
thoracic  lymph  in  beinp  more  cloudy,  often  l)loo<ly.  contains  more  solids,  has  a 
liipher  molecular  concentration  with  decreased  electrical  conductivity  (Jappelli 
and  d'Errico,  Zeit.  f.  Biol.,  1!)07  (.'SO)  1),  all  of  whicli  llndiii<rs  are  in  afjreement 
with  the  hypothesis  that  postmortem  lymi>h  flow  dejiends  upcm  ehanpes  in  the 
cells,  caused  liy  as))hyxia  and  not    dissimilar  <o  the  clianjres  of  acute  ne]dnitis. 


THE  CAU^EH  <>E  EDEMA  347 

feet  the  eolloitl-s  in  a  way  similar  to  acids,  or  tlirougli  the  conversion 
of  colloids  M^hieh  have  but  little  affinity  for  water  into  such  as  have 
a  fjfreater  affinity,  must  also  be  bonie  in  mind."  In  support  of  this 
theory  he  advances  evidence  which  lie  interprets  as  indicatin*^  that: 
(1)  "xVn  abnormal  production  or  accumulation  of  acids,  or  condi- 
tions predisposing  thereto,  exist  in  all  states  in  which  we  encounter 
the  development  of  an  edema.  (2)  The  development  of  an  edema  in 
tissues  is  antagonized  by  the  same  substances  which  decrease  the 
affinity  of  the  (hydrophilic)  enuilsion  colloids  for  water  (salts)  and  is 
unaffected  by  the  presence  of  substances  which  do  not  do  this  (non- 
■eleetrol5i;es).  (3)  Any  chemical  means  by  which  we  render  possible 
the  abnormal  production  or  accumulation  of  acids  in  the  tissues  is 
accompanied  by  an  edema." 

There  are  many  features  of  lymph  formation  and  edema  with 
which  this  theory  seems  to  harmonize  well,  and  others  with  w'hich  it 
•does  not  seem  to  agree  so  well,  if  at  all,  so  that  at  this  time  it  is  a 
fair  statement  that  the  theory  is  under  consideration,  but  the  limita- 
tions of  its  applicability  have  not  yet  been  agreed  upon.  It  has  met 
with  much  adverse  criticism,  some  of  w^hich  was  poorly  founded,  but 
the  fact  cannot  be  disputed  that  the  amount  of  water  that  colloids 
will  hold  varies  greatly  with  changes  in  the  colloids.  "We  may  not 
T\now  absolutely,  at  present,  whether  the  changes  that  take  place  in 
the  colloids  during  life  are  great  enough  to  alter  tlieir  water  content 
appreciably,  but  it  is  highly  probable  that  they  are.  In  many  in- 
stances the  principles  of  colloidal  hydration  offer  the  best  explanation 
■of  observed  conditions,  and  their  application  often  elucidates  matters 
more  satisfactorily  than  any  other  working  hypothesis.  Certainly 
they  cannot  be  disregarded  in  considering  the  factors  that  may  come 
into  iilay  in  producing  edema. 

Summary. — We  find  that  a  number  of  factors  may  be  considered 
as  responsible  for  edema,  some  of  them  being  prominent  in  one  in- 
stance, some  in  another,  but  in  feiv  cases  can  we  consider  one  factor 
alone  as  the  sole  cause.  In  most  of  the  forms  of  edema,  such  as  those 
due  to  renal  disease  and  cardiac  disease,  it  now  seems  probable  that 
either  osmotic  pressure  changes  or  changes  in  the  affinity  of  the  tissue 
<3olloids  for  water,  play  the  most  important  part ;  whereas  in  inflamma-\ 
tory  edema  there  can  be  no  question  that  alteration  in  the  capillary 
■walls  is  the  most  essential  factor.  But  the  mechanical  factor  of  blood 
pressure  cannot  be  disregarded,  although  by  itself  seldom  sufficient  to 
cause  edema ;  associated  with  other  factors  it  is  undoubtedly  an  im- 
portant agency,  for  there  are  few  edemas  that  are  not  associated  with 
increased  blood  pressure.  Hydremia  and  hydremic  ])lethora  may  al- 
most he  disregarded,  except  in  so  far  as  they  may  cause  altered  metab- 
olism in  the  tissues,  injuiy  to  vessel-walls,  over-saturation  of  the  blood 
■colloids,  and  decreased  osmotic  pressure  within  the  vessels.     Lymph- 


348  EDEMA 

atic  obstruction  is  possibly  a  factor  of  some  secondary  importance  if 
we  consider  that  distended  vessels  and  tense  tissues  may  occlude  the 
lymph  capillaries. 

SPECIAL  CAUSES  OF  EDEMA 

We  may  now  consider  which  of  the  above  factors  are  at  work  in 
bringing  about  edema  under  the  conditions  in  which  it  is  usuallj^ 
observed  clinically.  Before  taking  up  the  detailed  consideration  of 
edematous  conditions,  however,  it  may  be  well  to  call  attention  to  the 
fact  that  our  knowledge  of  edema,  and  especially  its  clinical  recog- 
nition and  study,  has  been  handicapped  by  the  lack  of  a  suitable  ob- 
jective method  of  detecting  and  measuring  edema.  We  are  in  the 
same  position  in  respect  to  edema  that  we  were  to  blood  pressure 
when  the  only  clinical  measure  was  the  clinician's  forefinger.  An 
attempt  to  remedy  this  defect  has  been  made  by  Schade,^"''  whose 
"elastometer"  reveals  and  measures  degrees  of  edema  not  discernible 
by  the  palpating  finger.  A  study  of  edema  with  this  instrument  in  the 
hands  of  Schwartz  ^°^  has  revealed  many  interesting  facts,  but  as  yet 
the  apparatus  is  too  complicated  for  general  clinical  use. 

"Cardiac"  Edema. — Passive  congestion  introduces  nearly  all  these 
factors,  for  in  addition  to  the  increased  blood  pressure  there  is  also 
an  opportunity  for  changes  in  the  capillary  wall,  either  from  stretch- 
ing and  thinning  of  the  cells  and  cement  substances,  or  from  "loss 
of  tone"  in  the  endothelium  surrounding  the  stomata  (Meltzer), 
or  from  toxic  injury  by  accumulated  products  of  tissue  metabolism. 
AVhen  the  stasis  is  nearly  complete,  or  if  it  is  complete  for  a  time  and 
then  relieved,  the  endothelium  may  be  injured  through  lack  of 
nourishment.  As  the  edematous  fluid  in  chronic  passive  congestion 
is  usually  of  a  watery  type,  poor  in  proteins,  the  edema  is  prob- 
ably less  dependent  upon  capillary  permeability  than  upon  other 
factors,  except  in  the  case  of  acute  stasis,  when  the  fluid  partakes  of 
the  character  of  the  exudates.  Presumably  the  accumulation  of 
crystalloids  within  the  tissues  also  plays  a  part  in  this  form  of  edema, 
as  the  osmotic  pressure  is  raised  in  tissues  having  deficient  oxygen 
supply.  But  Fischer  holds  that  the  reduction  in  oxidation  acts  chiefly 
by  increased  production  of  acids,  which  greatly  increase  the  affinity 
of  the  tissue  colloids  for  water  and  at  the  same  time  alter  the  colloidal 
state  of  the  capillary  endothelium  so  that  the  capillaries  become  more 
permeable.  Finally,  there  is  probably  more  or  less  obstruction  to 
lymphatic  outflow  because  of  the  increased  pressure  on  the  lymphatic 
channels,  and  perhaps,  also,  in  the  case  of  cardiac  incompetence,  ob- 
struction to  the  discharge  of  lymph  from  the  thoracic  duet  into  the 
sul)clavian  vein  against  the  higli  intravenous  ])i-essnre. 

Renal  Edema. — AVe  must  recognize  under  tliis  heading  two  dififer- 

soaZeit.  cxp.  Path.  u.  Thor.,  1!)]2    (11),  .309. 
30b  Arch.  Int.  Med.,   1910    (17),  396  and  459. 


SPECIAL  CAUSES  OF  EDEMA  349 

ent  types  of  edema.  In  acute  nephritis  {e.  g.,  in  scarlatina)  toxic 
materials  appear  to  be  the  chief  cause,  and,  as  Senator  contends,  in- 
jure alike  the  capillaries  of  the  renal  glomerules  and  of  the  sub- 
cutaneous tissues;  in  each  case  there  results  an  increased  permeability 
which  is  manifested  by  albuminuria  as  a  result  of  the  injury  to  the 
renal  capillaries,  and  by  edema  as  a  result  of  the  injury  to  the  tissue 
capillaries.  This  sort  of  edema  is  allied  to  that  produced  by  peptone 
and  similar  lymphag-o<iues,  and  we  might  well  imagine  that  the 
mechanism  consisted  merely  in  an  injury  to  the  capillaries  through 
which  excessive  fluid  is  driven  by  the  blood  pressure,  were  it  not  for 
such  observations  as  those  of  ]\Iendel  and  Hooker,-^^  who  found  that 
postmortem  flow  is  increased  by  these  lymphagogues  also.  We  can 
hardly  account  for  the  force  exhibited  in  postmortem  lymph  flow  on 
any  other  ground  than  that  it  is  furnished  b}^  osmotic  pressure  or 
colloidal  absorption,  unless  we  wish  to  fall  back  upon  "vital  activity" 
of  the  surviving  cells.  Hence  it  is  probable  that  even  in  the  edemas 
of  toxic  conditions,  such  as  acute  nephritis,  physico-chemical  factors 
play  a  part,  the  responsible  substances  probably  being  abnormal  or 
excessive  metabolic  products  of  the  cells  atit'ected  by  the  poisons.  An 
interesting  observation  made  by  Bence  ^-  is  that  nephrectomized  rabbits 
develop  an  edema  even  when  they  are  given  no  water  at  all ;  this  would 
seem  to  indicate  an  increased  affinity  of  the  tissues  for  water  when 
the  renal  functions  are  deficient.  Hydremia  is  always  a  favoring 
factor,  however,  and  probably  important  in  nephritic  edema,^^  while 
nearly  all  students  of  acute  experimental  nephritis  find  evidence  that 
the  resulting  edema  depends  very  much  upon  the  changes  in  the  vessel- 
walls. ^"^ 

In  the  more  common  edema  of  chronic  nephritis  we  have  to  con- 
sider, among  other  factors,  the  blood  pressure.  That  this  is  not  an 
essential  or  even  important  cause,  however,  is  shown  by  the  fact 
that  edema  is  usually  much  less  marked  in  interstitial  nephritis  with 
high  blood  pressure  than  it  is  in  parenchymatous  nephritis  with  a 
much  lower  pressure.  Toxic  substances  are,  of  course,  also  present  in 
the  blood,  and  may  alter  capillary  permeability ;  these  toxic  substances 
may  account  for  the  localized  edemas  and  erythemas  sometimes  ob- 
served in  nephritis.  But  jirobably  most  important  is  the  action  of 
the  crystalloids  which  the  kidney  does  not  excrete,  and  which  seem 
to  be  stored  up  in  the  tissues,  where  they  cause  transudation  of  water 
under  the  influence  of  their  osmotic  pressure.  For  example,  Rzent- 
kowski  ^■'  found  that  the  average  lowering  of  the  freezing-point  by  the 
eden:atous  fluid  in  nephritis  was  0.583°,  in  cardiac  dropsy  it  was  0.548°, 

31  Amer.  Jour,  of  Phvsiol.,  inn2    (7),  380. 
32Zeit.  f.  klin.  ]\red.,'inon    (67).  0(1. 
33Pearce,  Arch.  Int.  Med.,  1909   (.3).  422. 

34  See  Schmidt  and   Schlaver,  Deut.  Arch.  klin.  Med..   1911    (104).  44:    Tollak, 
Wien.  klin.  Woch.,   1914    (27).  98. 
35Berl.  klin.  Woch.,   1904    (41).  227. 


350  EDEMA 

and  in  tuben-iilous  jileuritis  0.526°.  This  indicates  that  the  osmotic 
concentration  of  the  fiuid  is  highest  in  renal  dropsy,  and  supports  the 
belief  that  here  and  in  cardiac  dropsy  osmotic  pressure  plays  a  more 
important  part  than  it  does  in  inflannnatory  exudation.  Of  the 
crystalloids  that  cause  accumulation  of  fiuid  in  the  tissues,  sodium 
i-hloride  seems  to  be  the  most  important. 

Retention  of  Chlorides  in  Edema. :^'' — From  the  investigations  made  l).v  numer- 
ous clinicians,  especially  the  French,  it  appears  that — (1)  in  nephritis  with 
edema  a  retention  of  sodium  chloride  frequently  occurs;  (2)  that  elimination 
of  the  clilorides  is  often  increased  durino;  periods  of  improvement  of  the  edema: 
(3)  that  a  reduction  of  the  amount  of  clilorides  in  the  diet  sometimes  causes  a 
great  improvement  in  the  edema,  while  administration  of  chlorides  may  make 
the  edema  mucli  worse.  There  are,  however,  observations  that  also  indicate  that 
chloride  retention  does  not  account  for  many  cases  of  renal  dropsy,  for  com- 
monly the  above-mentioned  conditions  are  not  fulfilled. st^a.  Nevertheless,  it  cannot 
be  denied  that  chloride  retention  is  sometimes  an  important  causative  factor  in 
the  edema  of  parenchymatous  nephritis. 37  If  the  retained  chlorides  obeyed  the 
ordinary  laws  of  diffusion,  we  should  expect  them  to  become  distriluited  alike  in 
the  bIoi)d  and  tissues,  so  that  they  would  merely  cause  an  equal  increase  in  the 
lluids  of  the  blood  and  of  t!ie  tissues;  that  is  to  say,  tliere  would  be  an  hydremic 
plethora  due  to  retention  of  water  in  the  body  by  the  accinnulating  chlorides. 
But,  according  to  a  number  of  observers,  there  is  a  specific  retention  in  tlie 
tissues,  which  Strauss  calls  "historetention,"  and  which  explains  the  local  edema. 
The  way  in  which  the  historetention  is  produced  is,  however,  not  understood,  and 
not  all  observers  accept  this  hypotliesis.  If  chlorides  do  bear  a  causative  rela- 
tion to  edema,  the  predilection  of  the  sulicutaneous  tissues  for  edematous  accumu- 
lations may  be  explained  by  the  observation  that  when  salt  is  given  to  an  animal 
an  undue  proportion  (28-77  per  cent.)  accumulates  in  the  skin.sTa  In  many 
conditions  other  than  nepliritis,  there  is  also  a  cliloride  retention  {e.  g.,  pneu- 
monia, cardiac  incompetence,  sepsis,  typlioid),  and  the  edemas  observed  in  these 
diseases  may  possibly  depend  upon  cliloride  retention,  as  many  Frencli  authors 
suggest.  Rumpf,  indeed,  often  found  more  chlorides  in  edematous  lluids  of  non- 
nephritic  origin  than  in  nephritic  edema.s'^b  Fischer  holds  that  the  retention 
of  chlorides  in  edema  is  secondary  and  not  primary,  for  he  foimd  that  tissues 
made  to  take  up  more  water  through  acidification,  also  take  up  an  increased 
amount  of  chlorides. 

'^  Inflammatory  Edema. — Although  here  the  alterations  in  the  cap- 
illary walls  play  an  essential  role,  as  shown  by  the  protein-rich  na- 
ture of  the  exudates,  yet  most  of  the  other  factors  are  added.  In- 
creased blood  pressure  is  prominent ;  lymph  outflow  is  impeded  by 
plugging  of  the  lymphatic  channels  by  clots  and  leucocytes,  and  by 
pressure  on  the  outside ;  there  is,  undoubtedly,  an  excessive  forma- 
tion of  metabolic  products  in  the  tissues,  to  cause  exosmosis,  and  the 

3«  Literature,  resumf"  bv  \Yidal  and  Javal,  Jour.  Phvsiol.  et  Pathol.,  1003  (5), 
1107  and  1123;  Rumpf,  Miinch.  med.  Woch.,  1005  (52),  303.  Review  in  Albu 
and  Neuberg's  "Mineralstoffweclisei,"  Berlin.  IflOO,  pp.  171-178;  Georgopulus. 
Zeit.  klin.  Med.,  !!)()()  (tlO),  411;  Cliristian,  Boston  Med.  and  Surg.  Jour.,  1008 
(158),  416;    Palmer,  Arcli.  Int.  Med..    I'M")    (15),  329. 

3Ga  See  Blooker,  Deut.  Arcli.  kliii.  y\vd..  liM)!!  ( Od ) .  SO;  Fischer,  "(iMlcnia  and 
Nephritis." 

37  See  Borcbardt,  Deut.  med.  Woch.,  1012   (38),  1723. 

37a  Scliade.  Zeit.  exp.  Patli.  u.  'Ilier..  1013  (14),  1.  Also  gives  an  interesting 
discussion  of  the  relation  of  the  skin  to  edema. 

37b  Breitmann  ( Zentr.  inn.  Med.,  1013  (34),  033)  describes  under  tlic  name  of 
"soda  dropsy"  a  form  of  edema  which  results  from  excessive  administration  of 
sodium  bicarl)onate  to  correct  acidosis  in   diabetes. 


NEUROPATHIC  KDKMA  351 

aspliyxial  conditions  in  intlaiiuHl  tissues  favor  acid  formation  wliichy 
may  cause  in  the  colloids  an  increased  affinity  for  water.  According 
to  Oswald  •'''''  the  pormoability  of  tlie  vi^ssels  for  proteins  becomes  spe- 
cifically altered  in  inflammation,  so  that  not  only  the  less  viscous 
albumin  and  pseudoglobulin  pass  throug'h  their  walls,  but  also  the  more 
viscous  euglobulin  and  fibrinogen.  To  this  class  of  edemas  belong 
also  the  urticarias  which  follow  the  ingestion  of  various  toxic  sub- 
stances, many  of  wliieh  can  be  shown  experimentally  to  be  lympha- 
gogues.  A  good  example  is  the  urticaria  which  often  follows  the  in- 
jection of  antitoxic  or  other  foreign  serums,  particularly  their  re- 
peated injection ;  in  experimental  animals  such  a  serum  may  cause 
death  very  quickly  by  acute  pulmonary  edema.  All  these  poisons 
probably  produce  urticarial  edema  by  injury  to  the  capillary  walls  in 
the  subcutaneous  tissues,  and  possibly  changes  in  the  hydrophilie 
properties  of  the  tissue  colloids  are  also  produced  by  the  poisons.  In 
the  action  of  vesicants  especially,  it  may  well  be  questioned  if  changes 
in  the  capillary  walls  and  active  hyperemia  are  not  supplemented  by 
local  metabolic  alterations.  The  edema  which  follows  the  sting  of 
insects,  which  are  known  to  secrete  into  the  wound  such  acids  as 
formic,  seems  to  be  a  particularly'  good  illustration  of  the  production 
of  edema  by  the  influence  of  acids  on  the  tissues  (Fischer). 

Neuropathic  Edema. — Until  we  understand  better  than  we  now 
do  the  manner  in  which  nervous  impulses  modify  metabolism,  it  will 
be  difficult  to  estimate  properly  the  importance  of  nervous  impulses 
in  the  production  of  edema.  That  nervous  control  is  a  possible  factor 
is  well  shown  by  many  experiments;  for  example,  simple  ligation  of 
the  femoral  vein  in  animals  does  not  cause  edema,  but  if  the  sciatic 
nerve  is  cut  the  vasoconstrictors  are  paralyzed,  and  edema  may  follow 
(Ranvier).^'-  In  this  case  the  nervous  influence  is  only  indirect 
through  its  vasomotor  effects.  Similarly,  stimulation  of  vasodilator 
fibers  may  cause  edema.  It  is  furthermore  possible  that  nervous 
stimulation  may  lead  to  excessive  metabolic  activity,  with  an  ac- 
cumulation of  crystalloidal  products  and  acids  sufficient  to  cause 
edema  when  supplemented  by  active  congestion  and  some  resulting 
pressure  upon  the  lymph-vessels.  There  are  certainly  many  instances 
in  which  edema  seems  to  depend  upon  nervous  disturbance ;  for  ex- 
ample, edema  in  the  area  of  distribution  of  a  neuralgic  nei've;  sudden 
joint  effusions  in  tabetic  arthropathy ;  and  especially  the  typical 
"angioneurotic"  edema. ^-"  The  only  explanation  that  seems  open  is 
the  one  given  above,  namely,  a  combination  of  local  hyperemia  and  in- 
creased metabolic  activity.     Even  the  urticarias  of  apparently  me- 

37c  A.  Oswald,  Zeit.  cxp.  Path.,  1010   (8),  226. 

3S  Similarly,  pulmonary  edema  follows  experimental  hydremia  onlv  when  the 
yapfi  are  cut  "(F.  Kraus,  Zeit.  exp.  Path..  1013  ( 14) ,  402) .    " 

3Sa  iletabolism  in  anfrioneurotic  edema  is  discussed  by  Miller  and  Pepper,  Arch. 
Int.  Med.,  1916    (18),  551. 


352 


EDEMA 


chanical  ori^n  {urticaria  factitia),  show  evidence  of  a  toxic  action, 
in  that  there  occurs  a  severe  nuclear  fragmentation  ( Gilchrist). ^^'^ 

Hereditary  Edema. — In  a  number  of  families  there  has  been  observed 
a  peculiar  inherited  tendency  to  the  occurrence  of  acute  attacks  of 
local  edema,  which  not  infrequently  have  proved  fatal  when  involving 
the  glottis.^^'=  There  can  be  little  question  that  these  instances  of 
hereditary  edema  depend  upon  a  nervous  affection  of  some  kind,  it 
being  practically  an  angioneurotic  edema ;  but  how  the  edema  is  pro- 
duced, and  wliat  the  nature  of  the  nei*vous  alteration  may  be,  are  as 
mysterious  as  are  most  other  so-called  "nervous  inheritances."  There 
also  are  cases  of  congenital  edema,  which  may  occur  repeatedly  in 
the  fetuses  of  the  same  mother  and  cause  habitual  miscarriage ;  ^^  and 
still  another  class  of  cases  in  which  the  children  are  born  apparently 
healthy,  but  develop  fatal  dropsy  when  a  few  weeks  old,***  Nothing 
is  known  as  to  the  cause  of  this  condition.  Patein  "  has  analyzed  the 
fluid  in  a  case  of  congenital  ascites  and  found  it  somewhat  more  like 
an  exudate  than  a  transudate. 

COMPOSITION  OF  EDEMATOUS  FLUIDS  ^- 

As  is  well  known,  the  composition  of  edematous  fluids  varies  greatly 
according  to  the  cause  of  the  edema  and  the  place  where  it  occurs. 
In  general,  non-inflammatory  edemas  (transudates)  contain  much  less 
protein  than  do  the  inflammatory  exudates,  as  is  shown  by  the  follow- 
ing table  of  analyses  by  Halliburton  ^^  and  by  Bernheim  's  **  deter- 
minations of  proteins  in  ascitic  fluids. 


Table  I 


Sp.gr. 

Parts  per  100  of  fluid 

Total 
protein 

Fibrin 

Serum- 
globulin 

3.002 

1.2406 

1.76 

Serunr 
albumin 

Acute  pleurisy 

1.023 
1.020 
1.020 

5.123 

3.4371 

5.2018 

0.016 
0.0171 
.0.1088 

2.114 

1.1895 

3.330 

Hydrothorax      .  "1 
Aver,  of  3  cases  J  ' 

1.014 

1.7748 

0.0086 

0.6137 

1.1557 

ssbJolins  Hopkins  Hosp.  Bull.,  1008   (19),  49. 

3SC  Literatu-re,  see   Fairbanks,   Amer.  Jour.  Med.  Sci.,   1904    (127).   Si 
and  French,  Quart.  Jour.  Med.,  1908    (1),  312. 

:ioW.   Fischer,  Berl.  klin.  Woch.,   1912    (49),  2403. 

40  Kd<ic\vorth,  Lancet.  1911    (181),  21(>. 

41  .Tour.  Pharm.  et  Chim.,   1910    (102),  209. 

••-  Many  data  are  jriven  l)v  Cerliartz,  llandhuch  der  Hiocheinie,  190S,  11 

43  Adarni,  Allbutfs  System,  1896    (1),  97. 

44  Quoted  by  Hammarsten,  "Physiological  Chemistry." 


Hope 


2),  137, 


COMI'OS/'noX  OF  EDKMATOl  s  j'l.iins 
Table  II 


353 


Ascitic  fluid  in 

Parts  of  protein  to  1000  c.  v'.  fluid 

Max. 

Min. 

Mean 

Cirrhosis  of  the  liver 

Bright's  disease 

34.5 
Ki.ll 

5.6 

10.10 

0.69-21.06 
15.6  -10.36 

Tuborculosis  and  idiopatliic  peritonitis 
Carpinomatous  peritonitis 

55.8 
54.20 

1S.72 
27.00 

30.7-37.95 
35.1-58.96 

Tlie  specific  g-ravity  varies  nearly  in  direct  proportion  to  the  amount 
of  proteins,  that  of  traiisndates  nsnally  being  below  1.015,  and  exu- 
dates above  1.018,  although  there  are  many  exceptions.  Indeed,  it  is 
often  very  difficult  to  decide  whether  a  given  fluid  is  an  exudate 
or  a  transudate.^''  According  to  Rzentkowski,*"  the  transudates  at 
the  moment  they  pass  out  of  the  vessels  are  simply  solutions  of  crystal- 
loids in  water  and  quite  free  from  protein ;  the  small  amount  of  protein 
found  in  transudates  he  ascribes  to  protein  pre-existing  in  the  tissue- 
spaces.  This  idea  is  hardly  acceptable  in  view  of  the  known  per- 
meability' of  the  vessel-walls  for  proteins  in  normal  conditions ;  more 
probably  in  cardiac  and  renal  dropsies  the  quantity  of  protein  escap- 
ing from  the  vessels  is  not  greatly  different  from  normal,  but  the 
excessive  fluid  escaping  in  these  conditions  carries  with  it  no  addi- 
tional proteins,  and  to  this  extent  transudates  in  statu  nascendi  are 
protein-free. 

Transudates,  even  when  produced  by  the  same  cause,  vary  in  com- 
position in  different  parts  of  the  body,  presumably  because  of  varia- 
tions in  the  permeability  of  the  vessels  in  different  vascular  areas;  just 
as  pleural,  pericardial,  peritoneal,  and  meningeal  fluids  normally 
differ  from  one  another.     Thus  C.  S.  Schmidt  *'  found  the  composition 

4s  Rivalta  (Eif.  Med.,  1903:  Bioehem.  Centr.,  1904  (2),  529)  has  siiirtrested 
the  following  test  to  distinguish  exudates  and  transudates:  Into  a  beaker  con- 
taining 200  o.c.  of  water  with  4  drops  of  glacial  acetic  acid,  let  fall  a  few  drojjs 
of  the  fluid  to  be  tested.  If  an  exudate,  a  bluish-white  line  is  left  transiently 
behind  the  sinking  drops,  due  to  precipitation  of  the  euglobulin  and  filjrinoyen. 
Tliis  test,  and  also  certain  modifications  (see  Rivalta.  Policlinico,  1910  (17). 
(i7fl),  seem  to  give  quite  reliable  results.  (See  I'jihard,  Berl.  klin.  Woch.,  1914 
(51),  1112.  With  tuberculous  effusions  Rivalta's  test  is  positive,  but  not  Mo- 
relli's  test,  which  consists  in  dropping  tlie  fluid  into  saturated  HgClo  solution,  a 
yellowisii  ring  of  all)uminate  forming  with  non-tuber:'ulous  exudates,  and  a  gran- 
ular precipitate  with  transudates.  (See  Zannini,  Gaz.  degli  Osped.,  1914  (4), 
461).  ]\Iemnii  (Clin.  Med.  Ital.,  1905,  No.  3)  suggests  the  larfrer  content  of 
lipase  as  a  means  of  distinction  of  exudates.  Tedeschi  (Caz.  degli  Osped..  1905 
(26),  SS)  states  that  e<jg-albumen  fed  in  large  amounts  appears  in  transudates 
and  not  in  exudates,  and  can  be  detected  by  the  biological  precipitin  test.  Sugar 
is  found  more  often  in  transudates    (Sittig). 

^o  Virchow's  Arch.,  1905    (179),  405. 

47  Hoppe-Sevler's  Phvsiol.  Chemie. 
23 


354 


EDEMA 


of  the  transudates  in  different  parts  of  the  body  of  a  patient  who  died 
of  nephritis  to  have  the  foHowing  composition : 


Table  III 


Pleural 

Peritoneal 

Subarachnoid 

Subcutaneous 

Water 

Solids 

Organic   matter 
Inorganic  matter     . 

963.95 

36.05 

28.i50 

7.55 

978.91 

21.09 

11.32 

9.77 

983.54 

16.46 

7.98 

8.48 

988.70 

11.30 

3.60 

7.70 

As  in  this  case,  the  general  iiile  is  that  while  the  proportion  of 
salts  remains  nearly  constant,  the  proportion  of  protein  in  edematous 
fluids  in  different  localities  varies  in  decreasing  order  as  follows : 
(1)  pleura;  (2)  peritoneum;  (3)  cerebrospinal;  (4)  subcutaneous.*"* 
In  the  last-named  location  the  specific  gravity  of  edematous  fluids 
may  be  as  low  as  1.005,  and  the  proteins  even  less  than  0.1  per  cent. 
(Hoffmann*'').  An  increase  in  solids  occurs  after  the  effusion  has 
existed  for  some  time,  presumably  because  of  absorption  of  water  and 
salts,  leaving  a  slowly  increasing  proportion  of  proteins.  Further- 
more, the  composition  of  the  patient's  blood  has  considerable  influ- 
ence on  the  composition  of  the  effusion;  this  is  particularly  true 
in  the  case  of  ascites  from  portal  obstruction,  the  contents  of  the  blood 
coming  from  the  intestine  during  digestion  modifying  the  composition 
of  the  ascitic  fluid.  Thus  IMiiller,"'**  in  a  case  of  portal  vein  throm- 
bosis, found  in  the  ascitic  fluid  of  a  patient  on  an  ordinary  mixed 
diet,  0.179  per  cent,  nitrogen;  on  a  protein-rich  diet,  0.2494  per  cent. 
N ;  on  a  protein-poor  diet,  0.1764  per  cent.  N.  In  cachectic  conditions 
the  proportion  of  proteins  is  less  than  in  stronger  individuals,  and, 
as  in  the  blood  plasma,  the  albumin  decreases  more  rapidly  than  the 
globulin  as  the  cachexia  advances  (Umber). ''^ 

Physical  Chemistry  of  Edema  Fluids. — The  differences  between 
transudates  and  exudates  depend  almost  solely  on  their  protein 
contents,  for  the  non-protein  elements  are  almost  identical  with 
iioi-inal  lymph  and  blood-serum,  which  naturally  must  be  so  since 
any  original  or  temporary  deviation  in  osmotic  pressure  must  be 
rapidly  e(|ualized  by  diffusion.  Thus  Bodon  ''-  finds  the  concentra- 
tion of  the  electrolytes  nearly  constant  in  spite  of  considerable  dif- 
ferences in  composition  of  various  edema  fluids,  indicating  that  the 
serosa  permits  passage  of  inorganic  salts  always  in  the  same  con- 

48  Javal  (Jour.  phys.  et  path..  1911  (13),  508)  places  Ihc  i\\\n\»  in  tliis  order: 
serum,  peritoneal,  pleural.  su1)cutaniH)us,  cerebrospinal. 

40  Deut.  Arch.  klin.  Alcd..   1SS9    (44),  313. 

r.o  Dent.  Arcli.  klin.  Med.,   1903    (76),  563. 

51  Zeit.  klin.  Med.,  1903    (48),  364. 

f'SPfliiper's  Arch.,  1904  (104),  519;  also  see  GaU'otti,  l.o  Sperimentale,  1901 
(55),  425. 


C(>]fl'<)S/T/(>\   or  KI>K\I.\T()V8  FLUIDS 


355 


centration,  while  lioldiii*--  back  the  organic  substances.  Transiidatfis 
contain  an  excess  of  NaCl  over  other  electrolytes,  while  in  exudates 
the  proportion  of  electrolytes  other  than  chlorides  is  increased  over 
the  finding:s  in  transudates.^^  The  surface  tension  of  exudates  is  lower 
than  that  of  transudates,'^  depending  chiefly  upon  the  glol)ulin  con- 
tent. Rzentkowski "'"'  found  some  slight  differences  in  molecular  con- 
centration as  indicated  by  the  freezing-point :  in  tuberculous  pleurisy 
the  average  lowering  was  0.523°,  that  of  the  serum  being  — 0.56°; 
in  cardiac  dropsy  the  subcutaneous  fluid  gave  — 0.548°,  and  in  renal 
dropsy  — 0.583°  ;  tuberculous  peritonitis,  — 0.523°  ;  cirrhosis  — 0.536°  ; 
carcinomatous  edema  — 0.547°.  Of  these  figures,  the  most  significant 
is  the  comparatively  high  molecular  concentration  of  the  fluid  in 
nephritis,  supporting  the  contention  that  the  cause  of  renal  edema 
is  retention  of  crystalloids.'"  Tieken  '"'  has  found  the  results  in 
transudates,  exudates,  and  other  body  fluids  shown  in  Table  IV. 


Table  IV 


Freezing- 

Freezing- 

Nature  of  Fluid 

Sp.  gr. 

point  of 
effusion, 
— °  C. 

point  of 
blood, 
— °  C. 

Disease 

Pleuritic   effusion    . 

1.016 

— 0..55 

—0.56 

Pneumonia,  lobar. 

((                     a 

1.018 

—0.55 

—0.55 

it                it 

"                     "     .        . 

1.018 

—0.54 

—0.56 

Tuberculosis. 

a                      li 

1.020 

—0.55 

—0.56 

It 

"                      "     .         . 

1,016 

—0.55 

—0.56 

it 

a                     It 

1.018 

—0.64 

—0.56 

Valvular  heart  disease. 

It                    It 

1.0.30 

—0.60 

—0.58 

Empyema ;   cyanosis. 

Pericardial     "   . 

1.018 

—0.55 

—0.56 

Pericarditis. 

It               it 

1.016 

—0.56 

—0.56 

" 

It               it 

1.012 

—0.56 

—0.56 

Hydropericardium. 

Ascitic  fluid 

1.024 

—0.60 

—0.56 

Cirrhosis  of  liver. 

te            it 

1.020 

—0.57 

—0.56 

a             a          (.' 

<C                  it 

1.018 

—0.58 

—0.56 

Tuberculous  peritonitis. 

it            it 

1.013 

—0.62 

—0.56 

Organic  heart  disease. 

C(                    It 

1,035 

—0.65 

—0.58 

General  peritonitis. 

Hydrocele  fluid 

1.016 

—0.56 

—0.56 

Tuberculosis. 

Cerebrospinal  fluid 

1.018 

—0.62 

—0.58 

Uremic  coma. 

«               .1 

1.016 

—0.64 

—0.68 

it               a 

tt               it 

1,020 

—0.64 

—0.64 

<(               it 

«               it 

1.014 

—0.56 

—0.56 

Tuberculous  meningitis. 

U                            it 

1.017 

—0.56 

—0.56 

Epidemic  meningitis. 

«                 it 

—0.56 

—0.56 

((                 it 

The  very  high  figures  for  effusions  in  nephritis  and  cardiac  incom- 
petence indicate  the  concentration  of  crystalloids  in  these  fluids,  and 

53Gruner,  Biocliem.  Jour.,  1907   (2),  383. 
54Trevisan,  Zeit.  exp.  Path..  1011    (10),   141. 
55Loc.  cit.,iG  and  also  Berl.  klin.  Woch.,  1904   (41),  227. 

56  Purulent  exudates  may  show  a  high  molecular  concentration    (-0.84°   in  one 
case),  due  to  decomposition  of  the  proteins  into  crvstalloids    (Rzentkowski). 
5' Amer.  Medicine,  1905   (10),  822. 


356  EDEMA 

support  the  belief  that  in  the  formation  of  both,  osmotic  pressure  is 
an  important  factor.'"'^ 

Edema  fluids  are  usually  alkaline  except  when  bacterial  changes 
lead  to  acid  formation,  but  they  are  always  able  to  neutralize  less  acid 
than  the  blood  of  the  same  individual  (Opie).  Bodon  ^'-  found,  how- 
ever, that  while  they  contain  alkali  that  can  be  neutralized  by  titration 
against  acids,  yet  they  resemble  the  blood  in  being  neutral  as  far  as 
the  presence  of  free  OH  ions  is  concerned. 

Protein  Contents. — As  indicated  in  the  tables  given  previously, 
these  vary  greatly  in  quantity  in  various  fluids ;  '''^  the  quantitative 
relations  of  the  different  varieties  of  proteins  have  been  less  studied. 
Serum-albumins  and  globulins  constitute  by  far  the  largest  part  of 
the  proteins,  fibrinogen  being  scanty  except  in  some  intiammatory 
exudates,  so  that  coagulation  very  seldom  occurs  spontaneously; 
pneumococcus  exudates  seem  particularly  rich  in  fibrinogen,  which 
coagulates  rapidly  and  firmly.  The  differences  in  the  proportion  of 
different  serum  proteins  in  transudates  is  attributed  by  A.  Oswald  ^^ 
to  the  relative  viscosity  of  these  proteins  .which  determines  their 
ability  to  pass  through  the  capillary  walls.  The  viscosity  of  serum 
proteins  varies  in  the  following  increasing  order:  albumin,  pseudo- 
globulin,  euglobulin  and  fibrinogen ;  heiice  in  transudates  w^e  may 
find  only  the  first  two,  or  perhaps  only  the  albumin,  while  in  exudates 
the  two  latter  appear.  Joachim ''-  found  in  pleural  transudates  and 
exudates  that  the  proportion  of  albumin,  euglobulin,  and  pseudo- 
globulin  is  always  lower  in  hydrothorax  than  in  pleurisy.  Of  dif- 
ferent forms  of  ascites,  the  largest  proportion  of  globulin  and  the 
smallest  of  albumin  occur  in  cirrhosis;  while  with  carcinoma  the  pro- 
portions are  reversed.  In  general  the  albumin  is  more  abundant  than 
the  globulin,"^  but,  as  Umber  ^^  has  found,  the  proportion  of  albumin 
sinks  more  rapidly  in  cachexia  than  does  the  globulin,  corresponding 
to  the  similar  changes  in  the  blood  proteins.  The  amount  of  protein 
lost  in  exudates  is  strikingly  shown  by  one  of  Umber's  cases  of  can- 
cerous ascites ;  during  one  year  the  fluid  removed  by  paracentesis  con- 
tained not  less  than  three  kilos  of  pure  j^rotein,  the  patient  weighing 
but  55.5  kilos. 

Several  authoi's  have  found  in  inflammatory  ascitic  exudates  a 
protein  having  ])hysica]  and  chemical  pi'opri'ties  much  resembling 
mucin;  it  has  Ix'cn  especially  studied  by  UiiilxT,'"  wlio  finds  it  quite 

■'■«  Meyer  and  His  (Deut.  Arch,  kliii.  Med.,  190,5  (S.")).  14!t)  olaiin  tliat  the  low- 
erinor  of  tlie  fr('ezing-j)oint  is  U^ss  tlian  tliat  of  tlie  1)1o(kI  in  exudates  wliile  form- 
ing, tlie  same  as  the  blood  wliile  stationary,  and  orreater  duiinji  absorption,  ^\llil■Il 
thev  consider  indicates  a  "vital  j)rooess"  on  the  part  of  tlie  cells. 

"o  Sec  also  v.  .Takscli,  Zeit.  klin.  .Med.,  1S!1;3   (2.*3) .  22;) ;  RzentkowsUi  ( Inc.  cit.)  4G. 

01  Zeit.  exp.  Path.,  IHIO   (8).  22(5. 

o--!  Pf!ii>j;er's  Arch.,   l!)(l.3    (!>:]),  .irjS. 

f'3  See  Epstein.  .Jour.  Kxp.  Med.,  1!)14    (20),  .3:U. 

<>•!  Zeit.  klin.  Med.,  1!»0:$  (48),  3(54:  also  llolst.  Tpsalalakar.  Forhand..  1!)04, 
p.  304. 


COMJ'OSIT/OX  OF  EDFAIATOU^  FLUIDS  357 

similar  Id  tiic  synovial  iiniciii  isolated  in  arthritis  by  Salkowski,  and 
calls  it  scrosaimicin. 

Non=Protein  Organic  Contents. — Proteoses,'^  leucine,  and  tyro- 
sine may  be  present  in  small  quantities  in  exudates,  being  produced 
by  autolysis'''"'  (Umber)  ;  and  also  mucoid  substances  (TTammarsten). 
Nucleoproteins  nuiy  be  ])resent  from  leucocytic  disintegration  in  exu- 
dates, as  well  as  the  products  of  their  further  splitting,  such  as 
purine's  and  phosphates,  (ilaldi  and  Appiani "'  found  uric  acid  con- 
stantly' in  amounts  between  0.0055  g.  and  0.0714  i:.,  in  all  exudates, 
of  which  seven  were  tuberculous  and  two  neoplastic.  In  three  trans- 
udates amounts  from  0.006  g.  to  0.011  g.  were  found.  Allantoin  is 
said  to  have  been  found  in  exudates  (Moscatelli),"*  but  this  is  doubtful. 

All  the  other  innumerable  components  of  plasma  may  be  found 
in  edematous  fluids;  thus  sugar'"''  and  urea  (Carriere)  ^*  are  often 
present,  as  well  as  other  extractives.  The  amount  of  urea  varies  quite 
as  it  does  in  the  blood  of  the  same  individual, ^^  and  it  seems  probable 
that  all  the  crystalloid  substances  present  in  the  blood  pass  freely 
into  and  from  inflammatory  exudates,  so  that  an  equilibrium  between 
]»lood  and  exudates  is  approximated.'-  Sugar  is  said  sometimes  to  be 
greater  in  amount  in  transudates  than  in  the  blood,  but  in  exudates 
it  is  usually,  if  not  always,  lower  than  0.1  per  eent.^**  Lecithin  is  al- 
ways present,  partly  bound  to  globulin  and  partly  free  (Christen)."^ 
Cholesterol  is  present  particularly  in  fluids  that  have  been  standing 
for  a  long  time  in  the  body,  appearing  often  as  visible  crystals  shining 
in  the  fluid  ;  it  probably  originates  from  degenerating  cells.  Glycogen 
is  not  present  (Carriere)."* 

Toxicity. — Contrary  to  earlier  ideas,  transudates  are  not  toxic, 
even  in  nephritis  (Baylac,"'  Boy-Teissier,"^  Laiforcade '"),  and  there- 
fore the  toxic  manifestations  frequently  observed  after  reduction  of 
edema  in  nephritis,  and  ascribed  to  absorption  of  poisons  in  the  trans- 
udates, are  probably  due  to  some  other  cause.     In  inflammatory  exu- 

osOpie,  Jour.  Exp.  Med.,  1007    (0),  391. 

66  Histidine  and  arginine  were  found  in  a  carcinomatous  exudate  hv  Wiener 
(Biochem.  Zeit.,  1912    (41),  149). 

67  Riforma  Med.,  1904,  p.  1.373:  also  Carriere,  Compt.  Rend.  Soc.  Biol.,  1899 
(51),  467. 

68  Zeit.  physiol.  Cliem.,  1899    (13),  202. 

69  Sugar  was  found  in  only  8  of  23  fluids  by  Sittig  (Biochem.  Zeit.,  1909  (21). 
14)  ;  but  is  present  in  pulmonary  edema  fluid  in  proportion  equal  to  or  even 
greater  than  the  blood    (Kleiner  and  !Meltzer). 

71  Javal  and  Adler,  Compt.  Rend.  Soc.  Biol.,  190G  (61),  235;  Roseiil)ero:,  Berl. 
klin.   Woch.,   1916    (53),   1314. 

72  Wells  and  Hedenburg,  Jour.  Infect.  Dis.,  1912  (11),  349;  Scheol,  Xord.  :\red. 
Laeg..  1916    (77),  610. 

70Hegler  and  Schumm.  :\red.  Klinik.   1913    (9),   1810. 

73  Cent.   f.   inn.   :\Ipd.,    1905    (26),   .329. 

74  Compt.  Rend.  Soc.  Biol.,  1899    (51),  467. 

75  Compt.  Rend.  Soc.  Biol.,   1901    (53),  519. 
-G  Ihid.,  1904   (56),  1119. 

77  Gaz.  heb.  Med.  et  Chir.,  Jan.  28,   1900. 


358  EDEMA 

dates,  of  course,  the  causative  agents  as  well  as  the  products  of  cell 
desti'uction  render  tlie  fluids  ])()is()nous. 

Enzymes  and  Immune  Bodies. — All  the  enzj'mes  of  the  plasma 
nuiy  appear  in  edematous  fluids,  being-  in  all  cases  probably  more 
abundant  in  exudates  than  in  transudates.  According  to  Carriere,'^ 
oxidases  are  inconstant,  even  in  exudates.  Lipase  is  said  to  be  much 
more  abundant  in  exudates  than  in  transudates."''  (Concerning  pro- 
teolytic enzymes  see  "Autolysis  of  Exudates,"  Chap,  iii.)  The 
various  immune  bodies,  cytotoxins,  hemolysins,  bacteriolysins,  ag- 
glutinins, etc.,  seem  to  pass  freely  into  both  transudates  and  exudates, 
and  their  presence  is  not  characteristic  of  either,-"  but  as  a  rule  the 
proportion  is  much  higher  in  exudates. ^^  Peptid-splitting  enzymes 
are  usually  found  in  such  fluids,'*-  especially  tuberculous  exudates,^-'^ 
and  these  enzymes  seem  to  be  difi:'erent  from  both  erepsin  and  trypsin. 
Probably  this  type  of  enzyme  is  more  often  present  than  trypsin. 
Antitryptic  activity  is  usually  high,  unless  exhausted  by  the  presence 
of  much,  trypsin  from  cell-rich  exudates.  Purulent  fluids  are  usually 
poor  in  ojosonins ;  ®^  in  non-purulent  fluids  the  opsonin  content  varies 
with  the  amount  of  proteins."*^  Turpentine  exudates  may  sometimes 
be  more  strongly  bactericidal  than  the  serum  of  the  same  animal.*^ 
Exudates  usually  contain  about  as  much  complement  as  the  serum, 
but  in  suppuration  the  complement  disappears ;  transudates  contain 
little  of  either  complement  or  hemolysins. ^^''^ 

Precipitin  Reactions,  etc. — Edematous  fluids  have  been  often 
used  as  a  source  of  material  in  immunizing  animals  against  human 
proteins.  The  precipitins  thus  formed  are  specific  for  human  serum 
or  for  the  proteins  of  the  effusion,  but  cannot  be  used  to  differentiate 
a  transudate  from  an  exudate,  or  a  hydrothorax  fluid  from  an  ascites 
fluid  (Quadrone ).'*''  Immune  bodies,  complement,  agglutinins,  and 
antitoxins  are  present  in  effusions ;  ^°  e.  g.,  the  common  use  of  blister 
fluid  for  the  Widal  test.  Furthermore,  according  to  Hamburger,*® 
edema  fluid  is  distinctly  more  bactericidal  than  normal  lymph. 

78Compt.  Kend.  Soo.  Biol.,  ISflO    (51),  oOl. 

TO  Zeri,  11  roliclinico,  1J10.3  (10),  No.  11;  :\lemmi,  ("lin.  :\lt'd.  Ital.,  ino.'>.  No.  3: 
Galletta,  Clin,  iiiwl.  Ital.,  1011    (50),  14.3. 

80  Granstriiiii,   Tnaug.   Dissort.,  St.   Petersburg,    lOO.i. 

81  Not  corroborated  by  Liidke,  Cent.  f.  Bakt.,  1907    (44).  2()S. 

82  Hall  and  Willianiscm.  .lour.  Path,  and  Bact.,  inil    (15),  .'551. 
82a  See  H.  Koeh,  Zeit.  Kinderhoilk..  1014    (10),   1. 

83  0pie.  .lour.  E.xper.  Med..  1!M)7    (0),  515. 

84  Brill  me,  Deut.  Areli.  klin.  Med..  1000   (96),  105. 
ssRastaedt,  Zeit.  Tnimunitiit.,  1012    (13),  421. 
85a  Aronstanim.  Cent.  f.  Bakt..  1014   (74),  32(1. 

80  Cent.  f.  Bakt.   (Ref.),  1905   (36),  270. 
88  Virohow's  Arch.,  1S09   (15(1).  320. 


VAh'lIJTIKH  OF  KDEMATOrx  FfJ  IDH  359 

VARIETIES  OF  EDEMATOUS  FLUIDS  -<:' 

On  tilt'  ])i-('c(Mliiiji-  ])ag<'s  liavc  been  mentioned  the  chief  differences 
in  the  characters  of  the  effusions  in  the  usual  sites,""  with  their  varia- 
tions in  protfMii  contents,  which  variation  agrees  with  Starling's  state- 
ment tliat  the  |)('rmeal)ility  of  the  capillary  wall  for  proteins  ditlt'ers 
normally  in  different  localities.  Some  of  the  other  ett'usion  fluids  not 
mentioned  jjreviously  have  particular  properties  of  some  interest. 

Subcutaneous  Effusions.'"^'' — When  of  non-inflammatory  origin  these 
are  very  watery,  having  ordinarily  a  protein  content  of  from  0.1  to 
0.2  gm.  per  100  c.c.  tliere  being  more  globulin  in  nei)hritic  than  in 
cardiac  dropsy.  The  non-coagulable  nitrogen  and  chloride  content 
are  not  so  high  as  in  the  blood  of  the  same  patients,  but  the  ash  is  the 
same  as  that  of  the  serum.  The  specific  gravity  may  be  as  low  as 
1.00."),  but  the  solids  increase  with  the  duration  of  the  edema. 

Hydrocele  and  Spermatocele  Fluids. — These  have  been  studied 
particularly  by  Hammarsten,  who  found  the  average  results  of  analyses 
of  seventeen  hydrocele  fluids  and  four  spermatocele  fluids  as  follows: 

Table  V 

Hydrocele  Spermatocele 

Water          9,38.85                           986.8.3 

Solids 61.15                            13.17 

Fibrin           0.59                              

Globulin 13.25                               0.59 

Seralbumin 35.94                               1.82 

Ether-extractive  bodies          .      .  4.02  1 

Soluble   salts 8.60  \-                       10.76 

Insoluble    salts 0.66  J 

Marchetti  ^^  found  in  ten  specimens  of  hydrocele  fluid  rather  higher  results  for 
the  solids  than  did  Hammarsten.  He  found  57.8  to  104.2  p.  m.  of  solids,  contain- 
ing organic  substances  48.8  to  95.02,  and  inorganic  substances  8.10  to  9.56;  pro- 
teins, 33.5  to  90.19;  ratio  of  globulin  to  albumin  as  2.56  to  9.11.  Among  the 
proteins  is  found  1  to  4  p.  m.  that  is  not  precipitated  by  heat.  Corresponding 
with  the  analytic  results,  tlie  specific  gravity  of  hydrocele  fluid  is  higher.  l.Olti  to 
1.026  as  against  1.006  to  1.010  for  spermatocele  fluid.  Cholesterol  is  often  abun- 
dant in  hydrocele  fluids,  appearing  to  the  naked  eye  as  glistening  scales. 
Patein  ^^  found  sugar  in  most  specimens  of  hydrocele.  Apparently  hydrocele  fluid 
stands  intermediate  in  properties  between  transudates  and  exudates. -'3 

Meningeal  Effusions.-'* — Normal  meningeal  fluid  differs  from  all 
other  serons  fluids  in  being  clear  and  Avatery,  in  its  low  specific  gravity 
(1.004  to  1.007),  in  containing  but  a  trace  of  protein  which  is  chiefly 

80  Chemistry  of  Pus  and  Sputum  are  discussed  under  Inflammation,  Chapter  x. 

90  Literature  and  resume  on  pleuritic  exudates,  see  Ott,  Chem.  Pathol,  der 
Tuberc,  1903,  p.  392. 

noaSee  Epstein,  Jour.  Exper.  Med.,  1914    (20).  334. 

91  Lo  Sperimentale,  1902    (56).  297. 

92  Jour,  pharm.  et  chim.,  190(i  (23),  239;  also  Conipt.  Rend.  Soc.  Piol.,  1906 
(60),  303. 

93  Vecchi,  Gaz.  Med.  Ital.,  1912  (63  1,  211;  Epstein,  Jour.  Exp.  Med.,  1914  (20), 
344. 

94  Resume  by  Blumenthal,  Ergeb.  der  Physiol.,  1902  (1),  285;  Blatters  and 
Lederer,  Jour.Amer.  Med.  Assoc,  1913   (60)j  811, 


360  EDEMA 

globulin  (with  a  trace  of  proteose  ( ?)),  and  0.05-0.13  per  cent,  of  a 
reducing-  substance  that  is  proba])ly  <«-lucose,''^'  which  is  decreased  in 
acute  suppurative  meningeal  intlannnation  (Jacob).'"'  Halliburton 
gives  the  following  analyses  of  pathological  accumulations  of  such 
fluids : 

Table  VI.      (Spina  bifida.) 

Case  1  Case  2  Case  3 

Water         989.75  989.877  991.6.5S 

Solids         10.25  10.123  8.342 

Proteins  0.S42  1.(502  0.199 

Salts  )        .      .      .      .  f.f..^f.  (    0.631  3.02S 

Extractives     i        .      .      .      .  '^-     "  *    7.890  5.115 

The  percentage  of  solids  in  spina  bifida  is  thus  a  little  higher  than 
in  normal  meningeal  fluids.  In  hydrocephalus  the  percentage  of  solids 
is  rather  greater,  as  seen  in  Table  VII. 

T.\.DLE  VII.      (Hydrocephalus.) 

Case  1                 Case  2  Case  3 

Water 986.78              984.59  980.77 

Solids 13.22                15.41  19.23 

Proteins  and  extractives    .      .          3.74                  6.49  11.35 

Salts          9.48                  8.92  7.88 

Normal  cerebrospinal  fluid  seems  to  be  hypertonic  to  the  serum  of 
the  same  animal,**'  and  slightly  more  alkaline  than  the  blood.**^  In 
meningitis  the  alkalinity  is  often  lowered.''*''  According  to  Fuchs  and 
Kosenthal,-*"  the  average  freezing-point  of  the  cerebrospinal  fluid  is 
loW'Cred  about  the  same  in  all  diseases  (A  = — 0.52°  to  — 0.54°)  ex- 
cept in  tuberculous  meningitis,  wdiere  it  is  much  less  (average  — 0.43°). 
The  amount  of  potassium  is  about  the  same  as  in  the  blood/  and  not 
increased  in  degenerative  diseases  of  the  central  nervous  system ;  ^°- 
after  death  the  amount  is  much  increased  by  post-mortem  changes. 
Calcium  is  almost  constant  at  5  mg.  per  100  c.c,  or  about  one-half  as 
much  as  in  the  plasma.^"  In  diseases  associated  with  destruction  of 
brain  tissue,  such  as  general  paralysis  and  epilepsy,  choline  or  some 
other  base  -  may  be  found  in  the  spinal  fluid.  (See  "Choline,"  Chap. 
iv.)  Under  pathological  conditions  the  amount  of  protein  varies 
greatly  and  to  some  extent  characteristically.  Thus,  in  syphilis  the 
euglobulin    is  so   greatly-    increased   that   it   is   readily   identified   by 

95  Schloss    and    Schroeder,    Amer.    Jour.    Dis.    Child..    191(1     (11),    1;    Hopkins, 
Amer.  .Jour.  Med.  Sci.,  1915    (150),  847. 
no  Brit.  :Mcd.  .lour.,  1912,  Oct.  26. 

07  Ravaut,  Presse  nied.,  1900  (8),  128;  Zanier,  Cent.  f.  Phvsiol.,  1896   (10),  353. 
98Hur\vitz  and  Tranter,  Arch.  Int.  Med.,  1916   (17),  828.  ' 
n«aLevinson,  Arch.  Pediatri'-^.   1916    (33),  241. 
onWien.  med.  Presse,  1904    (4.H.  2081   and  2135. 

1  Mvers,  Jour.  Biol.  Cheni..   PHi'.l    (6),   115.  literiilure. 

la  Pvosenhloom  and   Andrews.    Arch.    Int.   ^Med.,    1914    (14),   536. 
ih  llalvcrson  and   IJerfrciin,  .lour.  Biol.  Cli.'in.,  1917    (29),  337. 

2  Kaufmann,  Zeit.  piiyniol.  ("hem.,  1910  IM)),  343;  Lai,i:nel-Liivastiiu>  and 
Lasusse,  Compt.  Rend.  Soc.   Biol..   1910    (OS),  803. 


\.\ini:Tii:s  or  j:i>i:\i  \t<>(  s  fij  ids  :^61 

various  |)i'('('i|)ilat  ioii  met  liods,  ■  while  in  more  acute  iuHaiumatious 
libriuoficii  appears.'  Acc()rdiii<i-  to  .Mott  ^  the  fluid  is  especially  rich 
in  luicleiu  in  progressive  paralysis,  and  lipoids  are  increased  in  the 
fluid  in  degenerations  of  the  cent!-al  nervous  system.  Pathological 
fluids  show  also  s])ecific  alterations  in  their  colloidal  property  of 
])rev(>ntiiig  pi-eeipitation  of  colloidal  suspensions  by  electrolytes  (the 
"Goldzahl"  of  Zsigmondy)."  The  surface  tension  is  higher  than  that 
of  the  serum  and  is  not  characteristically  altered  in  disease.'""  The 
increased  organic  matter  of  pathological  fluids  raises  the  permanganate 
reduction  index.'"' 

Cholesterol  can  be  found  in  all  cases  of  mental  disease,  the  amount 
not  bearing  any  relation  to  the  type  of  psychosis  (Weston)  ;  ^  ordinar- 
ily 0.2  to  0.7  mg.  per  100  c.c.  is  found.  The  changes  in  P2O-,  content 
in  disease  are  doubtful,**  while  the  amount  of  reducing  substances  is 
said  to  be  increased  in  disease.-'  In  general  the  inflammatoiy  fluids 
in  the  spinal  canal  resemble  exudates  elsewhere,  but  usually  the  con- 
centration of  the  different  components  is  relatively  low,  except  the 
chlorides.^"  Normal  cerebrospinal  fluid  contains  no  antiprotease  (for 
leucoprotease),  as  does  the  fluid  in  many  cases  of  chronic  inflamma- 
tions; in  acute  inflammation  proteases  may  appear  (Dochez  ^^).  Pep- 
tid-splitting  enzymes  are  especially  abundant  in  meningitis. ^^'^  Anti- 
bodies pass  from  the  serum  into  the  cerebrospinal  fluid  only  in  minimal 
amounts  or  not  at  all,  except  when  inflannnatory  exudation  occurs, 
and  even  then  the  antibody  concefitration  is  usually  low,^-  and  even 
simple  chemicals  enter  the  normal  spinal  fluid  but  very  little, ^^  ex- 
cept perhaps  alcohol.^*  According  to  Rosenbloom  "-''^  there  is  no  crea- 
tin  or  creatinine.  It  contains  normally  from  2  to  4  mg.  of  amino-N. 
per  100  c.c,  or  about  half  that  in  the  blood,  without  definite  changes 
in  syphilis.^'"'  There  is  almost  the  same  amount  of  urea  as  in  the 
serum  of  the  same  person,  i.  e.,  20  to  42  mg.  per  100  c.c.^*'=  Substances 
giving  the  ninhydrin  test  appear  in  meningitis,'^''  but  Rosenberg  states 

3  See  Xogiiclii,  Jour.  Exp.  Med.,  inon    (11),  604. 

4  See  :\Iestrczat.,  Rev.  d.  Med.,  ]!)10,  )).  ISO;  KafYka.  Dciit.  nied.  \Voch.,  1013 
(39),  1874. 

5  Lancet.  July  0,  1910. 

6Lange,  Zeit."  Cliemother.,  1012  (1),  44;  Spiit,  Zeit.  Tmnniiiitat.,  I'll.i   (23),  426. 
eaKisch  and  Kemcrtz.  IMiinch  Med.  Woch.,   1914    (20),   1097. 
6b  See  Hoffmann  and  Schwartz,  Arch.  Int.  Med.,  1916    (17),  293. 

7  Jour.  Med.  Res.,   1915    (33),   119. 

8  Apelt  and  Schumm.  Arch.  Psvchiat  u.  Xervenkr.,   190S    (44),  84.5. 
9,]»acob,  Brit.  Med.  Jour.,  Oct. '26,  1912. 

10  Java!,  Jour.  pbys.  ct  ])atli.  gen..  1911    (15),  508. 

11  Jour.   Exp.   Med.,    1909    (11),   718. 

iia  Major  and  Nobel,  Arch.  Int.  Med.,  1914    (14),  383. 

isLeniaire  and  Debre,  Jour,  physiol.  et  patli.  gen.,  1911    (13),  233. 

13  See  Eotkv,  Zeit.  klin.  :Med.,  "l912    (75),  494. 

14  Schottmiiller   and    Scliumm,   Neurol.   Zbl.,    1912    (31),    1020. 
i4aRiochem.  Bull.,  1916    (5),  22. 

14b  Ellis,  et  al..  Jour.  Amer.  Med.  Assoc,  1915    (04).  126. 
14c  Ellis  and  Cullen,  Jour.  Biol.  Chem.,  1915   (20),  511. 
i4dNol}el,  jMiinch.  med.  Woch.,  1915    (62),  1355,  1786. 


362  EDEMA 

that  even  with  the  hig'liest  indieanemia '^"^  no  indicaii  is  found  in  the 
spinal  fluid.  Su<iar  is  present  in  from  0.07  to  0.085  per  cent,  and  is 
not  modified  sig'uificantly  in  mental  diseases.^^'"  Tliere  is  only  a  very 
small  amount  of  diastase,  not  bearing  any  constant  relation  to  the  cell 
count. ^^^ 

Wound  secretions  obtained  from  lar<'P  aseptie  wounds,  mostly  amputation 
stumps,  liavc  been  studied  by  Lieblein.i'''  Tlie  reaction  is  generally  alkaline, 
jrlobulin  and  alb\imin  abundant,  but  tibrinoaen  scanty,  total  nitr()ii:en  beiu'i  less 
than  that  of  tlie  blood  and  decreasino-  from  day  to  day;  the  proportion  of  albumin 
increases  and  trloluilin  decreases  as  liealino:  ])rou:resses.  Occasionally  albumoses 
were  found,  but  only  (m  the  first  day  in  asejjtic  wounds:  if  found  later,  thev 
ffenerallv  were  antecedent  to  suppuration  (concerning  suppuration  see  ''Inflam- 
mation,"' (hap.  X.) . 

Blister  fluid  is  generally  rich  in  solids  and  proteins  (40-65  p.  m.).  In  a  burn 
blister  i^b'irner  i"  found  50.31  p.m.  proteins,  among  which  were  11.50  p.  m.  globulin 
and  but  0.11  p.  m.  fibrin;  also  a  substance  reducing  copper  oxide,  but  no  pyro- 
catechin.  By  refractometric  determinations  the  amount  of  protein  in  blister 
fluids  is  in  "dirwt  pro])ortion  to  tlie  amount  in  the  blood. i"  Antiljodies  of  all 
sorts  seem  to  pass  readily  into  blister  fluids,is  although  tlie  complement-fixation 
reaction  is  not  so  strong  as  with  the  blood. isa 

Chylous  Effusions.^' — Fat  may  be  present  in  effusions  in  sufficient 
quantity  to  cause  a  milky  appearance,  either  from  escape  of  chyle 
from  a  ruptured  or  obstructed  thoracic  duct,  or  throuoh  fatty  degen- 
eration of  the  cells  in  the  effusion  or  the  lining  of  the  walls  of  the 
cavity.  The  former  are  designated  as  chylous,  the  others  as  chyli- 
foDii  or  adipose  fluids,  but  it  is  ^lot  always  easy  to  distinguish  be- 
tween them.  The  composition  of  the  fluids  in  true  chylous  exudates 
will  vary  according  to  the  food  taken  and  the  amount  of  fat  the  food 
contains,  and  will  resemble  the  composition  of  chyle,  except  to  the 
extent  that  it  is  modified  by  the  effusion  or  absorption  going  on  in  the 
cavity.  They  are  characterized  by  strong  bactericidal  powers  as  evi- 
denced by  lack  of  putrefaction  after  long  standing. 

Analyses  of  human  chyle  are  scanty.  Panzer  20  found  00.29-04.5.3  per  cent, 
water ;' 5.47-0.71  per  cent,  solids;  0.80-1.04  per  cent,  inoraanic  salts;  2.1(5  per 
cent,  coagulable  protein;  0.50  per  cent,  ether-soluble  material;  also  diastatic  en- 
zyme, soaps,  and  occasionally  traces  of  cholesterol,  lecithin,  and  sugar.  Carlier.-i 
in  a  specimen  from  a  child,  obtained  very  similar  results,  excejit  tluit  tlie  salts 
were  much  less  a])undant.  The  proteins  and  fats  vary  greatly  witli  the  did  :  thus 
Sollmann  --  found  variations  in  the  jiroteins  from   1.S5  to  (i.o  per  cent. 

i4eBerl.  kl.  Wocli..  1016   (53),  1314. 

i4f  Weston,  Jcnir.  Aled.  I'es..  lOKi  (35),  100;  Kraus  and  Corneille.  .Tour.  Lab. 
Clin.  ^Fed.,  1016    (1),  685. 

i-*«  lycsclike  and  Pincussohn.   Deu(.  nicd.  Woclis.,   1017    (43),  S. 

ir.  T?eit..  klin.   C'liir.,    1002    (35),   43. 

Ki  Ilammarsten,   .Amer.  ed.,   1004.   p.  224. 

17  En  gel  and  Ors/ag,  Zeit.  klin.  :Med.,  1000   (67),  175. 

i«  p:isenberg.   Dent.    med.   Woch.,    1000    (35),    (il3. 

isaBusclike  and  Zimmcrmann.  Med.  Klinik,  1013    (0),  1082. 

in  Literature  bv  (iandin,  Krgeb.  inn.  ]\1e<l.,  1013    (12),  218. 

20  Zeit.   phvsiol.  Chem..    1000    (30),    113. 

21  British  'iMed.    Jour..    1002     (ii),    175. 

22  Amer.  .Tour.  Ph\sinl.,  10(17  (17).  4S7  :  see  also  Ilainill.  Jour.  Pliysiol..  1006 
(35),  151. 


VARIETIES  OF  EDEMATOl  S  I'LL  IDS  363 

p](l\\ai'(ls  ■■  found  but  60  definitely  established  cases  of  cliylous  or 
cliyliforni  ascites  in  the  literature  up  to  1895;  and  of  31  indisputable 
cases  studied  at  autopsy,  in  21  there  was  established  the  existence  of 
a  ruj)tui'e  in  the  thoracic  duct  or  lacteals.  Boston-*  in  1905  was 
able  to  collect  126  cases,  includin*?  both  chylous  and  chyliform  as- 
cites, and  notes  an  associated  fosinophilUi  in  a  case  studied  by  him. 
Chylous  ascites  tluid  often,  but  not  always,  contains  sugar,-'^  but 
it  may  disappear  after  havinj>-  once  been  present ;  the  amount  of  fat 
is  small,  usuall\'  about  1  per  cent.,  and  the  fluid  is  rich  in  solids. 
If  due  to  a  ruptured  thoracic  duct,  it  may  ])e  ])()ssible  to  detect  special 
fats  taken  in  the  food,  e.  g.,  butter-fats  (Straus).-''  The  reaction  is 
usually  alkaline  or  neuti-al.  and  some  specimens  coagulate  spontane- 
ously. Specific  gravity  varies  from  1.007  to  1.040,  the  average  being 
about  1.017.  Perhaps  the  most  important  characteristic  is  the  varia- 
tion produced  by  changes  in  diet.-'  Zdarek  -*  found  in  a  chyle-cyst 
2.7  per  cent,  of  fats,  7.2  per  cent,  of  proteins,  and  0.05  per  cent,  of 
sugar;  feeding  of  fats  increased  their  amount  in  the  cyst  and  star- 
vation decreased  it.  Schumm  -^  found  in  the  solids  of  such  a  cyst, 
35.76  per  cent,  of  fat,  some  of  which  was  in  the  form  of  calcium  soap. 

Chijlothorax  fluid  is,  of  course,  quite  similar  to  that  of  chylous 
ascites.  Thus.  Buchtala  ^"^  found  91.34  per  cent,  of  water,  8.66  per 
cent,  solid,  4.86  per  cent,  protein,  2.5  per  cent,  fat,  0.26  per  cent, 
cholesterol,  and  0.94  per  cent.  ash.  Similar  figiires  were  obtained  by 
Salkowski  "^  and  others. 

Chyluria,^^'^  which  seems  to  depend  upon  an  abnormal  connnunica- 
tion  between  the  lymphatics  of  the  receptaculum  chyli  and  the  kid- 
ney,^- shows  no  particular  chemical  features  beyond  those  of  an  ad- 
mixture of  a  considerable  amount  (100  to  1000  c.c.  per  day)  of  chyle 
with  the  urine.  Carter  ^^  found  the  amount  of  fat  in  the  urine  to  rise 
with  increase  of  fat  in  the  food.  In  some  cases  chyle  escapes  directly 
into  the  bladder  or  ureter  from  the  lymphatics,  in  others  the  fat  may 

23  ;Medicine.  1S95  (1),  257;  also  see  "Chem.  ii.  morph.  Eigensehaften  fett- 
haltiiie  Exsiulaten,"  St.  ^Mutermilch,  \Varscliau.  1003;  Comev  and  IMeKibhen, 
Boston  Med.  and  Surg.  Jour.,  1003   (14S),  100. 

24  .Jour.  Amer.  Med.  Assoc,  1905    (44),  513. 

25  For  example,  v.  Tabora  ( Deut.  nied.  Wocli.,  1004  I  30),  1505)  found  as  high 
as  0.864  per  cent,  of  sugar  in  a  tvpical  case. 

26  Arch.  Physiol,  et  Pathol..  1886' (Ser.  3,  vol.  8),  367. 

2T  A  sample  of  the  composition  of  1  liter  of  chylous  ascitic  fluid  is  shown  by 
the  analysis  in  the  case  studied  by  Comev  and  McKibben  Hoc.  cit.)  :  Specific 
gravity,  1.010;  solids,  21  gm. ;  protein,  0.75  gm.;  urea,  1.28  gm.:  fat,  1.45  gm. : 
inortranic  matter.  8  gm.;  peptone  (?)  and  sugar,  present;  fibrinogen,  mucin, 
nucleo-albumin.  and  uric  acid  absent. 

2SZeit.  f.  Heilk.,  1006    (27),  1. 

2n  Zeit.  phvsiol.  Chem.,  1006    (40),  266. 

soZeit.  phvsiol.  Chem.,  1010   (67),  42. 

3iVirchow"s  Arch..  1000  (108),  180;  also  Tulev  and  Graves,  Jour.  Amer.  :\Ied. 
Assoc.  1016    (66),  1844;   Patein,  Jour,  pharm.  Chim.,  1015    (11),  265. 

3ia  Review  of  literature  bv  Sanes  and  Kahn,  Arch.  Int.  iled.,   1016    (17),   181. 

32  See  Magnus-Lex-v.  Zeit.  klin.  Med.,  1008   (66),  482. 

33  Arch.  Int.  Med.,"  1916   (18),  541. 


364  EDEMA 

be  excreted  directly  from  the  blood,  independent  of  lymphatic  abnor- 
mality ;  in  some  cases  the  fluid  entering'  the  urine  is  true  chyle  and  in 
others  it  is  lymph. 

Ascites  adiposus  is  characterized  by  the  absence  of  sugar  and  by  a 
liigher  percentage  of  fat,  the  maximum  observed  being  6.4  per  cent. 
It  is  ascribed  to  fatty  metamorphosis  of  cells,  particularly  in  carcino- 
matous and  tuberculous  exudates;  Edwards  was  able  to  show  experi- 
mentally that  a  transudate  may  change  from  serous  to  cellular,  and 
later  come  to  contain  fat. 

Pseudochylous  effusions  are  also  observed,  not  onl}^  in  the  abdom- 
inal and  thoracic  cavities,  but  even  in  the  fluid  of  the  edematous  legs 
and  scrotum ;  these  resemble  chylous  fluids  in  being  turbid  or  milky, 
but  are  said  to  contain  little  or  no  fat.  The  turbidity  is  ascribed 
chiefly  to  lecithin,  which  is  largely  combined  with  the  pseudoglobulin 
of  the  fluid  ( Joachim). ^^  Possibly  in  some  cases  the  turbidity  is 
j)artly  or  largely  (Poljakotf)  ^''  due  to  poorly  dissolved  proteins. 
Strauss  ^"^  has  noted  the  occurrence  of  this  form  of  ascites  particu- 
larly in  chronic  parenchymatous  nephritis,  but  believes  the  turbidity 
has  a  local  origin.  Hammarsten  has  observed  a  similar  turbidit}^ 
due  to  mucoid  substances,  as  also  have  Gouraud  and  Corset.^'  The 
pseudo-chylous  effusions  have  a  lower  freezing  point,  a  lower  specific 
gravity,  lower  fat  and  greater  lecithin  content  than  typical  chylous 
ascites.  Gandin,^''  however,  questions  the  possibility  of  always  differ- 
entiating the  three  types  of  turbid  fluids  as  above  indicated.  Collect- 
ing all  the  recorded  analyses  in  the  literature  he  finds  wide  discrep- 
ancies, as  indicated  in  the  following  table:  (The  maximum  and  mini- 
mum percentage  figures  are  given  for  each  component  determined 
quantitatively,  with  the  average  in  parentheses.) 


Chylous 

Adipose 
(Chyliform) 

Pseudochylous 

Ether  extract 

O.O60-9.2    (1.65) 

0.1-4.3    (1.15) 

0.007-1.86    (0.25) 

Cholesterol 

+   in  7,     —  in  2 

+   in  4 

+  in  3,     —  in  2 

Lecithin 

+   in  4,     —  in   1 

+   in  3 

+   in  20,  —  in  2 

Sugar 

+   in  46,  —  in  28 

+   in   1,  —  in 

4 

-f-  in   15,  —  in   14 

Dry    residue 

;?.  1-10.6    (6.2) 

1.6-11.7    (5.1) 

1.2-7.6    (2.0) 

Protein 

(».!»-7.7    (;5.5) 

0.6-6.8    (3.0) 

0.1-4.2    (1.4) 

"Pepton."' 

+   in   (!,     —  in  4 

4-    in   1,  —  in 

2 

+   in   1,     —  in  5 

Asii 

0.1-1.0    (0.5!)) 

0.45-1.03    (O.6.- 

i") 

0.40-0.00    (0.73) 

It  is  quite  evident  that  although  the  pseudochylous  fluids  usually 
contain  little  fat,  they  often  contain  more  than  the  minimal  content 
found  in  the  other  forms.  P^ach  type  of  fluid  overlaps  the  others  in 
one  respect  or  anotiier.  Gandin  states  that  to  produce  a  turbid  fluid 
but  0.01-0.1  per  cent,  of  finely  emiilsionizcd  fat  is  necessary,  and  lie 

34Munch  nied.  W'odi.,  1003  (50),  1015;  also  thristen.  Cent.,  f.  inn.  Med..  1005 
(26),  320;  Wallis  and  ScJiolherj,',  Quart,  .lour.  Med.,  1010  (3),  301;   1011    (4).  153. 

35  Fortsclir.  d.  Med.,  1003  (21),  lOSl  ;  also  iiauslialter,  Coinj)!.  Heiid.  Soc  l!i(d., 
1910    (6H),  550. 

30  Note  to  Poljakoll's  article;  also  Hiochein.  Ceiitr.,   l!(i):{    (1).  437. 

37Conipt.  Rend.  Soc.  Biol.,  1006   (60),  23. 


VllEMISTh'Y  or  l'.\i:i   MOTHnUAX  365 

believes  tliat  milky  fluids  always  incaii  admixture  of  cliyle,  rejecting 
the  terms  pseudochylous  and  chyliform  as  unwarranted.  He  admits 
that  fluids  may  contain  droplets  of  fats  not  emulsionized,  and  hence 
not  millrs',  which  may  be  properly  called  adipose  fluids.  There  are  no 
characteristic  chemical  differences  in  tlie  fats  extracted  from  the  dif- 
ferent types  of  fluids. 

CHEMISTRY  OF  PNEUMOTHORAX 

Til  conneetioii  witli  the  subject  of  exudates  the  above  topic  iiuw 
apjn'opriately  be  considered.  The  composition  of  the  gases  found 
in  the  pleural  cavity  in  pneumothorax  will  necessarily  vary  greatly 
according  to  the  cause.  If  the  pleural  cavity  is  in  free  communica- 
tion with  the  exterior,  the  gas  will  be  simply  slightly  modified  air: 
for  example,  Ewald  ^^  found  the  following  proportions  in  the  gases 
in  such  a  pneumothorax :  CO.,  1.76  per  cent. ;  0,  18.93  per  cent. ; 
and  79.31  per  cent.  X.  Here  the  proportion  of  CO,  is  even  a  little 
less  than  in  ordinary  expired  air,  which  contains  3.3-3.5  per  cent. 
When  air  enters  a  closed  pleural  cavity  and  no  effusion  follows,  it  is 
slowly  absorbed  until  a  mixture  of  about  90  per  cent.  N,  4  per  cent.  0 
and  6  per  cent.  CO^  results ;  but  if  there  is  a  serous  effusion  the  oxygen 
disappears  nearly  or  quite  completely  (Tobiesen).^^  In  a  seropneu- 
mothorax Ewald  found.  8.13  per  cent,  of  CO2,  1.26  per  cent,  of  0,  and 
90.61  per  cent,  of  X,  which  is  quite  similar  to  the  proportions  of  the 
gases  in  dry  pneumothorax.  Purulent  pneumothorax  generally  shows 
more  COo  than  the  serous  form,  the  average  in  the  former  being 
15-20  per  cent.,  in  the  latter  7.5-11.5  per  cent.  The  average  of 
the  analyses  in  six  cases  of  pyopneumothorax  is  given  bj'  Ew^ald 
as  18.13  per  cent.  CO.,  2.6  per  cent.  0,  and  79.81  per  cent.  X'.  In 
open  pyopneumothorax  the  gas  approaches  more  closely  the  com- 
position of  air,  but  usually  shows  a  slight  excess  of  CO,;  it  is  thus 
possible  by  a  determination  of  the  carbon  dioxide  to  determine  quite 
accurately  whether  a  given  pneumothorax  is  in  communication  with 
the  outside  air.  The  transformation  of  a  purulent  into  a  putrid 
pneumothorax  is  accompanied  by  an  increase  of  COo,  even  as  high 
as  40  per  cent,  having  been  found.  The  products  of  decomposition 
by  the  putrefactive  saprophytes  also  are  present,  one  analysis  having 
shown  4.3  per  cent,  of  hydrogen.  6.25  per  cent,  of  methane,  and  traces 
cf  hydrogen  sulphide. 

Infection  of  a  pleural  effusion  by  gas-producing  organisms  may 
also  convert  it  into  a  pneumothorax,  although  this  is  not  a  common 
occurrence.  The  gases  then  present  are  the  same  as  the  organisms 
produce  in  similar  culture-media,  modified  somewhat  by  absorption. 
The  anaerobic  gas-producing  organisms  have  been  found  as  the  cause 

38  Complete  literature  and  resume  given  bv  Clemens,  in  Ott's  '"Cliem.  Patli.  der 
Tubereulose,"'  Berlin,  1008.  p.  4f)ti. 

30  Beitr.,  z.  Klin.  d.  Tuberk.,  1011  (1!)).  451;  1011  (21),  100;  Peut.  Arcb.  klin. 
Med.,  1914   (115),  399. 


366  EDEMA 

of  suuli  gaseous  accumulations;  it  is  questionable  if  the  ordinary  path- 
ogenic organisms  can  cause  a  pneumothorax,  since  they  are  for  the 
most  part  not  capable  of  producing  gas.  The  colon  bacillus  produces 
gas  in  sugar-containing  media,  but  the  amount  of  sugar  in  the  patho- 
logical exudates  is  too  small  to  yield  any  considerable  amount  of  gas; 
an  exception  is  the  pleural  eflfusion  in  diabetes,  and  pneumothorax 
from  infection  of  the  pleural  effusion  in  a  diabetic  by  B.  coli  has 
been  reported.  Complete  quantitative  analyses  of  the  gas  in  this  form 
of  pneumothorax  seem  not  to  have  been  made,  but  ^lay  found  about 
20  per  cent,  of  CO,.  The  combustibility  of  the  gas  has  frequently 
been  noted,  and  is  probably  due  to  hydrogen  and  methane. 


CHAPTER    XIII 

RETROGRESSIVE  CHANGES  (NECROSIS,  GANGRENE, 
RIGOR  MORTIS,  PARENCHYMATOUS  DEGENERA- 
TION) 

NECROSIS 

We  recognize  that  a  cell  is  alive  through  its  reproducing,  func- 
tioning, and  its  taking  on  and  utilizing  nutritive  substances;  yet 
at  tlie  same  time  we  appreciate  that  a  cell  may  do  none  of  these  things 
and  still  be  alive.  For  example,  a  bacterial  spore  is  quite  inert 
physically,  and  exhibits  no  chemical  activity,  yet  it  is  by  no  means 
dead,  since  it  still  possesses  the  latent  power  to  assume  again  an 
active  existence  under  suitable  conditions.  In  pathological  condi- 
tions we  are  accustomed  to  recognize  the  fact  that  a  cell  is  dead  by 
certain  alterations  in  its  structural  appearance,  particularly  disin- 
tegrative changes  in  the  nucleus ;  but  this  is  exactly  equivalent  to 
recognizing  that  an  animal  is  dead  by  the  appearance  of  postmortem 
decomposition,  for  most  of  the  characteristic  histological  changes  of 
necrosis  are  merely  postmortem  changes  in  the  cell.  A  cell  may  be 
dead  and  show  absolutely  none  of  these  microscopic  disintegrative 
changes,  either  because  it  has  not  been  dead  long  enough  for  them 
to  have  taken  place,  or  because  the  changes  have  been  prevented 
by  some  means,  just  as  we  can  prevent  the  appearance  of  postmor- 
tem decomposition  by  embalming.  For  example,  if  we  examine  mi- 
croscopically the  mucous  membrane  of  the  stomach  of  a  person  who 
has  died  immediately  after  taking  a  large  ([uantity  of  carbolic  acid, 
although  to  the  naked  eye  this  mucous  membrane  is  hard,  white, 
and  definitely  necrotic,  yet  we  find  the  histological  picture  presented 
by  the  cells  almost  absolutely  unchanged  from  the  normal.  The 
cells  are  dead,  but  they  have  been  so  "fixed"  that  postmortem 
changes  could  not  affect  their  structure.  All  cells  examined  by  ordi- 
nary histological  methods  are,  of  course,  dead — killed  by  the  fixing 
agents  outside  of  the  body,  in  the  same  way  that  the  carbolic  acid 
fixes  them  within  the  body.  It  is  evident,  therefore,  that  it  ma.v  be 
very  difficult  to  determine  always  whether  a  cell  is  dead  or  not.  Part 
of  the  difficulty,  perhaps,  lies  in  our  failure  to  appreciate  that  not  all 
parts  of  a  cell  die  at  the  same  time;  i.  e.,  the  causes  of  different  chem- 
ical processes  of  the  cell  reside  in  its  different  intracellular  enzjones, 
and  these  are  not  necessarily  destroyed   alike  by  the  same   agents. 

AVe  recognize  that  after  an  animal  is  dead  as  a  whole  the  various 

367 


368  RETROGRESSIVE    CHANGES 

cells  of  its  body  do  not  die  for  some  time,  as  shown  by  the  following 
examples:  (1)  We  can  cause  the  heart  to  beat  for  a  considerable 
period  after  its  removal  from  the  body;  (2)  if  we  perfuse  a  mixture 
of  g'lycocoll  and  benzoic  acid  throug-h  the  kidney  of  a  recently  killed 
animal,  synthesis  of  these  substances  into  hippuric  acid  will  occur; 
and  (3)  the  epithelium  of  the  skin  can  be  removed  from  the  body  of 
an  animal  lono:  after  death  and  transplanted  successfully  on  another 
animal.  So,  too,  in  ordinary  cell  death  (necrobiosis)  not  all  the 
enzymes  are  destroyed  together.  When  all  are  destroyed  at  once, 
as  by  strong  chemicals  or  by  heat,  the  customary  disintegrative 
changes  do  not  take  place.  If,  however,  not  all  the  enzymes  are 
thrown  out  of  function,  then  the  others  may  be  able  to  act,  producing 
the  disintegrative  changes  by  which  histologists  ordinarily  recognize 
cell  death.  These  tlisintegrative  changes  are,  for  the  most  part,  ap- 
parently brought  about  by  the  intracellular  proteases,  that  is,  through 
autolysis.  This  may  be  shown  as  follows:^  If  we  take  two  pieces 
of  fresh  normal  tissues  from  an  animal,  and  in  one  kill  the  enzymes 
by  heating  to  100°  C,  then  implant  both  aseptically  into  the  abdom- 
inal cavity  of  an  animal  of  the  same  species,  it  will  be  found  that 
the  changes  that  follow  in  the  two  will  be  very  unlike.  In  the  un- 
heated  tissue  nuclear  changes  soon  occur,  so  that  they  lose  their  ca- 
pacity for  taking  up  basic  stains,  the  cytoplasm  becomes  granular 
and  fragmented,  the  tissue  becomes  friable  so  that  it  is  difficult  to 
secure  good  sections,  and  the  changes  are  in  general  similar  to  those 
seen  in  areas  of  necrosis.  The  boiled  tissue,  on  the  other  hand, 
retains  its  capacity  for  nuclear  staining  for  months,  except  at  the 
periphery,  where  it  is  slowly  attacked  by  leucocytes  and  the  enzymes 
of  the  blood  plasma.  Therefore  it  would  seem  that  the  characteristic 
changes  of  necrosis  depend  ehietly  upon  the  intracellular  enzymes, 
rather  than  upon  the  infiltrating  plasma  as  Weigert  -  and  other  early 
writers  imagined.  In  areas  of  anemic  necrosis  (see  "Infarcts") 
we  have  another  case,  in  which  the  oxidizing  enzymes  are  thrown 
out  of  function  through  lack  of  oxygen,  while  the  otlier  enzymes  are, 
presumably,  at  first  luuiffected.  From  studies  of  infarcts  it  would 
seem  that  the  intracellular  proteases  bring  about  the  subsequent 
nuclear  and  cytoplasmic  alterations,  but  that  the  eventual  digestion 
of  the  area  is  accomplished  by  the  invading  leucocytes  working  slowly 
inward  from  the  periphery.  Apparently  when  the  sui^i)]y  of  materials 
from  outside  ceases,  and  when  the  oxidation  ]>ro('esses  of  the  cells  no 
longer  accomplish  necessary  steps  of  synthetic  reai-tions  or  destroy 
products  of  protein  catabolism,  the  proteases  continue  to  split  proteins 
without  the  balancing  by  the  above-mentioned  factors,  with  a  resulting 
disintegration  of  the  cells. 

Karyoljjsis  and   Ldri/orrhf.rls  are,  Ihcu,   Ihe   i-esult  of  an   autolytic 

1  Wells,  .Tour.  Med.  Kcsoarcli,  liMXI    (15  1.   lift. 

2  Cent.  f.  Path.,  1801    (2),  78,5. 


NECROSIS  369 

process,  whicli  is  perhaps  due  to  intracellular  proteases  that  act  spe- 
cifically on  nucleoproteins,  and  which  may  be  designated  as  nucleases.^ 
Nuclear  staining  by  the  usual  methods  depends  upon  an  affinity  of  tlie 
acid  nucleoproteins  (in  which  the  nucleic  acid  is  not  completely 
saturated  by  j)r()teins)  for  basic  dyes.  Presumably  in  karyolysis  the 
first  step  consists  in  a  splitting  of  the  nucleoprotein  of  the  chromatin 
into  nucleic  acid  and  protein;  this  can  be  accomplished,  according 
to  Sachs,  by  the  ordinary  trj'psin,  and  presumably,  therefore,  by  the 
trypsin-like  enzymes  of  the  cell.  Corresponding  with  this  cliange 
we  should  expect  the  free  nucleic  acid  to  give  an  intense  staining 
with  basic  stains,  and  this  has  frequently  been  described  by  those 
who  have  studied  the  cytological  changes  in  anemic  necrosis,*  and 
called  pijoiosis.  As  supporting  this  view  still  further  may  be  quoted 
Arnheim  's  "'  observation  that  in  alkaline  solutions  the  nucleus  soon 
stains  diffusely  and  weakly,  and  not  at  all  after  twelve  to  eighteen 
hours;  this  is  to  be  explained  by  the  fact  that  nucleic  acid  is  both 
dissolved  and  neutralized  by  alkaline  solutions.  Acids  developed  in 
injured  cells  may,  by  combining  with  the  basic  elements  of  the  n\\- 
cleoproteins,  render  them  still  more  acid  and  highly  basophilic ;  thus, 
in  muscles  showing  waxy  degeneration  from  accumulation  of  lactic 
acid  the  muscle  nuclei  will  be  found  pycnotic  (see  waxy  degenera- 
tion). After  the  nucleic  acid  has  been  freed  from  the  protein  by 
the  autolytic  enzymes,  it  is  still  further  decomposed  by  the  "nu- 
clease" or  similar  intracellular  enzymes  that  have  the  property  of 
splitting  nucleic  acid  into  the  purine  bases  that  compose  it — cor- 
responding with  this  change  the  hyperchromatic  nucleus  loses  its 
affinity  for  stains,  and  karyolysis  is  complete.  "When  extensive  ne- 
crosis occurs  there  will  result,  therefore,  an  increased  elimination 
of  purines,  as  was  found  by  Jackson  and  Pearce  ^  in  animals  with 
severe  hepatic  necrosis  from  hemotoxic  serum. 

A  careful  analytical  study  of  the  chanses  taking  place  in  the  autolyzinor  spleen, 
for  the  purpose  of  correlatino^  the  chemical  and  microscopical  chancres,  has  been 
made  by  Corper,"  whicli  corroborates  tlie  interpretation  of  necrosis  advaiu-e<l 
above.  Pie  found  tliat  during  the  stage  when  pycnosis  is  tlie  cliief  feature  tliere 
is  no  appreciable  change  in  the  nucleus;  tliat  is.  the  nucleic  acid  has  not  been 
split  into  free  purines  and  the  rest  of  its  components:  at  this  stage  but  I'ttle 
change  has  occurred  in  the  lecithin,  and  a  very  slight  amount  of  proteolysis  is 
demonstrable.  During  the  stage  of  karvorrhexis  and  karyolysis  the  most  active 
disintegration  is  taking  place,  about  one-fourth  of  the  nucleic  acid  becoming  dis- 
integrated by  the  time  all  nuclear  structures  have  disappeared :  in  tlie  same 
period  nearly  lialf  the  lecithin  (phosphatids)  is  hydrolyzed.  while  about  one- 
fourtli  the  coagulable  protein  has  l)een  hydrolyzed  into  non-coagulable  compcninds. 
After  this  stage  tlie  changes  are  very  slow.  It  is  somewhat  surprising  to  find 
that    wlien    no    vestige    of    nuclear    substance    remains    in    stainable    form,    tliere 

3  See  Purine  Metabolism,  Chap.  xxi. 

4  Schmaus  and  Albrecht,  Virchow's  Arch.,  1895  (138).  supp.,  p.  1;  Ergeb.  a\W. 
Pathol..  1806  (3).  486   Hiterature). 

■"'Virchow's  Arch..  ISnO    (120),  367.       • 
••Jour.  Exper.  Med.,  1007    (0),  560. 
TJour.  Exper.  Med.,   1912    (15),  429. 
24 


370  RETRoa  RJ^KSJ  1  y;  cii.w  a  eh 

still  rcniains  tlircc-foiutlis  of  the  nucleic  acid  in  an  intact  condition.  Corper 
publishes  a  scries  of  plates,  toficther  with  the  ciiemical  details,  thus  establishing 
a  standard  wliercby  tlie  histological  changes  can  be  interpreted  in  terms  of  tiie 
chemical  changes  which  cause  them. 

It  may  be  observed  that  autolysis  of  aseptically  prosevved  tissues 
outside  the  body  is  much  more  rapid  than  is  the  autolysis  of  infarcts. 
Mnd  similar  aseptic  necrotic  areas  within  the  body.  This  may  be  due 
to  either  or  both  of  two  factors :  '^  First,  autolysis  is  much  slower  in 
alkaline  than  in  acid  media;  outside  the  body  autolyzing  tissues  de- 
velop an  acid  reaction  which  favors  their  autolysis;  within  the  body 
this  is  checked  by  tlie  plasma.  Second,  the  ])'iasnia  contains  inhib- 
iting substances,  which  also  may  interfere  with  self-digestion  in  the 
body.  In  corroboration  of  the  above  may  be  recalled  the  fact  that 
large  necrotic  areas  show  autolysis  first  in  the  center,  where  the  alka- 
line, antagonistic  body  fluids  presumably  cause  the  least  effect.  Fur- 
thermore, it  has  been  found  by  Wells  ^  that  the  histological  changes 
of  autolysis  proceed  much  faster  in  tissues  placed  in  serum  that  has 
been  heated  to  destroy  the  antibodies  than  in  unheated  serum.  Leuco- 
cytes, as  Opie  has  shown,  contain  autolytic  enzymes  acting  best  in  an 
alkaline  medium,  hence  they  perform  their  digestive  function  readily 
at  the  periphery  of  necrotic  areas,  and  coagulated  tissue  proteins,  when 
acted  upon  by  body  fluids,  prodtice  chemotactic  substances  whidi  at- 
tract leucocytes  to  dead  areas."-' 

When  a  cell  dies,  certain  physical  changes  occur  that  are  probably 
of  considerable  importance.  The  permeability  of  the  cell  wall  is  al- 
most immediately  increased,  so  that  all  diffusible  substances  readily 
pass  through,  i.  e.,  its  semipermeable  character  is  lost.  This  we  see 
particularly  in  plant  cells,  which  lose  their  turgor  with  their  semi- 
permeability,  and  therefore  the  plant  wilts.  The  cell  structure  is 
also  disintegrated,  and  as  a  result  coordination  of  the  cell  chemistry 
is  at  once  destroyed.^"  Intracellular  enzymes  escape  into  the  blood 
from  areas  of  local  death  of  cells,^"''  or  as  an  agonal  manifestation  in 
general  death."*"  Various  dyes  which  cannot  penetrate  living  cells 
may  stain  dead  or  dying  cells."^  These  changes  depend  on  alterations 
ill  permeability,  and  as  permeability  determines  electrical  resistance. 
Osterhout  has  used  the  resistance  of  plant  cells  as  an  indicator  of 
vitality.  He  finds  that  normal  cells  have  a  rather  constant  resistance, 
which  is  reduced  by  anything  that  lowers  the  vitality  of  the  cell,  and 
in  direct  ])r()p()i'tion  to  the  degree  of  injury  or  loss  of  vitality.^'"'     The 

"Literature  and  more  complete  discussion  under  "Autolysis."' 

o.Tour.  Med.  Researcli,   }<)()(]    do),   149. 

naBiirger  and  Dold,  Zoit.  Tmmnnitiit..  1014   (21),  .378. 

10  See  V.  Prowazck.  Biol.  C-frM..  1!)00  (29),  291.  ri.ici  suggests  tli.it  in 
dead  proteins,  aldeliydes  and  amino  •adicals  iniite  witli  one  anotlier  to  form  cyclie 
compounds    (Arcli.  sci.  phys.  nat.,   191.')    (40),   ISl). 

i":i  Maiidelbaum,  ^liinch".  med.   Wooli.,    1914    (Ol).   4()1. 

I'lhSchiillz,  .Miinch.  med.  Woch.,  191.3    ((10),  2.'')12. 

IOC  Sec  Steckelmacher,  Beitr.  path.  Anat..  101.3   (.-)7),  314. 

10(1  See  Science,  1914    (40),  488. 


e.ir.sA'.v  OF  \i:(h'i)Sis  371 

temiK'raturc  I'oefficicnt  is  also  considerably  lower  in  dead  than  in  living 
tissne.'^  AVlien  seeondary  disintegrative  changes  occnr  in  the  proto- 
plasm, with  the  formation  of  many  small  molecules  from  the  large 
molecules  of  the  cell,  both  osmotic  pressure  and  electrical  conductiv- 
ity increase  rapidly.  Changes  in  the  permeabilit}^  of  cell  protoplasm, 
however,  may  be  of  considerable  degree  without  necessarily  indicating 
serious  injury  of  the  cells  (Osterhout).^"'' 

A  principle  of  colloid  chcmistrv,  the  alteration  of  colloids  with  time,  has  an 
interestinpr  bearing  on  liie  question  of  ajjing  and  natural  death  of  tissues. i-'  It 
is  cliaracteristic  of  colloidal  solutions  (\vliicli,  of  course,  is  what  cells  are),  tliat 
they  continuously  clianfj:e  in  tlieir  properties,  the  change  being  generally  in  the 
direction  of  aggregation  of  the  disperse  colloidal  particles,  with  a  resulting  ten- 
dency to  ])recipitatioii  or  coaguiatio!! :  the  gels  tend  to  decrease  in  elasticity  and 
U>  l)econie  moi-e  turbid,  associated  with  which  are  alterations  in  tlieir  per- 
meability to  crystalloids.  A  gelatin  mass  possesses  its  maximum  elasticity  tiiree 
or  four  hours  after  it  is  first  formed;  and  crystalloids  penetrate  fresh,  quickly- 
formed  gels  at  first  more  rapidly  than  later.  As  Bechhold  says,  we  can  inuigine 
(1)  a  relation  of  such  facts  to  the  greater  elasticity  of  young  tissues;  (2)  to  a 
))resumably  greater  permeability  for  crystalloids  and  hence  more  rapid  metab- 
olism: (3)  to  the  decreasing  water  of  tlie  tissue  with  age  (94  per  cent,  of  water 
in  the  fetus  of  three  months.  00-60  per  cent,  at  birth,  and  58  per  cent,  in  adults)  : 
(4)  to  the  demonstrated  greater  permeability  of  yoiuig  nerve  tissues  for  vital 
stains,  etc.  ''In  general  we  can  say  that  the  tissue  colloids  decrease  in  their 
water  attinity  (Qiiellharl-eit)  both  in  animal  organisms,  which  become  poorer  in 
water  with  age,  and  in  plants,  as  shown  by  the  hardening  of  older  j^lant  tissues.'' 
The  bearing  of  these  principles  on  the  problem  of  senility  and  degeneration  of 
elastic  tissue,  regeneration  and  many  other  subjects  is  obvious. 

CAUSES  OF  NECROSIS 
Anemia. — After  the  cutting  off  of  blood-supply,  cells  soon  undergo 
morphological  changes  that  we  recognize  as  indicating  tlieir  death, 
and  after  a  time  they  also  become  incapable  of  returning  to  their  nor- 
mal condition  when  the  blood-supply  is  re-established,  probably  be- 
cause of  these  structural  changes.  In  just  what  way  lack  of  nourish- 
ment causes  death  has  not  been  determined,  but,  as  has  been  before 
suggested,  it  seems  probable  that  it  is  because  catabolic  processes  are 
no  longer  balanced  by  anabolic  processes,  and  with  these  latter  oxi- 
dizing enzymes  seem  to  be  inseparably  associated,  as  far  as  our  pres- 
ent knowledge  shows  us.  That  the  loss  of  oxygen  alone,  with  other 
materials  presumably  supplied  to  the  cells  in  adequate  amount,  may 
cause  necrosis,  is  shown  by  the  presence  of  marked  hepatic  necrosis 
in  animals  kept  a  week  in  atmospheres  extremely  low  in  oxygen  (5-9 
per  cent.).^-^  The  nature  of  the  chemical  changes  taking  place  in  a 
cell  when  oxygen  is  deficient  must  be  very  different  from  the  normal 
changes,  and  hence  abnormal  toxic  substances  maj  accumulate,  e.  g., 
excessive  amounts  of  organic  acids.     Were  it  not  that  the  proteolytic 

11  Oaleotti's  earlier  observations  with  animal  tissues  (Zeit.  f.  Biol.,  100.3  (45). 
65)  do  not  harmonize  with  Osterhout's  results,  and  Galeotti's  idea  that  there  is  a 
special  degree  of  ionization  cliaracteristic  of  living  cells  is  not  established. 

loe  Botan.  Gaz.,  1015   (50),  242. 

12  See  H.  Bechhold,  "Die  Kolloide  in  Biologie  und  Medizin."  Dresden,  1012,  p.  65 
12a  Martin,  Bunting  and  Loevenhart,  Jour.  Pharmacol.,  Proc,  1916   (8),  112. 


372  RETROGRESSIVE    CHANGES 

enzymes  continue  in  action  after  nutrition  is  shut  off,  the  cells  might 
remain  in  a  completely  unaltered  condition  for  an  indefinite  period, 
and  capable  of  resuming-  their  function  when  nourishment  is  again 
supplied,  which  is  decidedly  contrary  to  the  facts.  (The  general 
features  of  anemic  necrosis  have  been  already  discussed  in  the  pre- 
ceding paragraphs,  and  also  under  the  subject  of  infarction.) 

Thermic  Alterations. — These  have  been  studied  particularly  in 
conneetion  with  the  cells  of  the  lower  organisms.^^  While  some  uni- 
cellular organisms  can  survive  a  temperature  of  69°,  most  of  them 
are  killed  at  from  40°-45°.  For  the  great  majority  of  metazoa  the 
nicfximum  temi)erature  lies  below  45°,  and  in  the  case  of  marine 
species  below  40°."  The  heating  is  accompanied  by  the  appearance 
of  granules  in  the  cytoplasm,  which  become  larger  until  the  condi- 
tion of  "heat  rigor"  sets  in.  Kiihue,  in  1864,  showed  that  in  muscle 
cells,  at  least,  there  is  contained  a  protein  which  becomes  turbid 
through  partial  coagulation  at  40°,  and  Halliburton  ^^  has  found 
that  in  nearly  all  tissues  are  globulins  coagulating  at  from  45°-50° ; 
it  is  probable,  therefore,  that  the  granules  formed  in  heated  cells  are 
produced  through  coagulation  of  these  proteins.  The  importance  of 
this  coagulation  in  determining  death  is  not  yet  fully  established, 
but  it  would  seem  to  be  very  great.  Halliburton  has  observed  that 
in  both  muscles  and  nerves  to  which  heat  is  applied,  contractions 
occur  at  various  temperatures,  corresponding  exactlj^  with  the  tem- 
peratures at  which  the  several  varieties  of  the  proteins  of  the  cells 
coagulate.  Furthermore,  Mott  ^"^  has  found  that  the  temperature 
that  is  immediately  fatal  to  mammals  (47°)  is  exactly  the  same  as 
the  coagulating  temperature  of  the  lowest  coagulating  protein  of 
nerve-cells.  This  fact  is  undoubtedly  of  great  practical  importance 
in  causing  death  from  fever,  for  although  47°  C.  (117°  F.)  is  prob- 
ably never  reached  in  man,  yet  application  of  much  lower  tempera- 
tures, even  42°  (108°  F.),  for  a  few  hours  will  cause  coagulation 
of  these  proteins  (all  proteins  coagulate  at  less  than  their  ordinary 
coagulation  point  if  the  heating  is  continued  for  a  long  time).  It 
would  seem  from  the  above  observation  that  heat  may  cause  cell  death 
through  coagulation  of  the  proteins.  AVhethei-  the  cell  death  is  in 
any  way  dependent  upon  destruction  of  the  enzymes  by  heat  has  not 
been  ascertained ;  but  as  most  enzymes  are  not  destroyed  much  be- 
low 60°-70°,  it  seems  improbable  that  they  are  greatly  injured  at 
the  temperatures  at  which  cells  are  killed.  It  is  possible,  however, 
that  under  the  conditions  in  which  enzymes  exist   in  the  cell   they 

i- Literature,  see  Davenport,  "Experimental  Aforphology,"  New  'Sork,  1S!)7; 
Schmaus  and  Albrecht,  Erpebnisse  der  Pathol..   1896    (,1,  Abt.   1),  470. 

14  The  adaptation  of  animal  cells  to  hipli  temperatures  is  an  interosliiii,^  topic, 
especially  in  view  of  sueli  results  as  tliose  of  Daliinfjer,  wiio,  by  raisiui:  tlie  teni- 
peraturo  gradually  durin<j  several  years,  caused  llaffeliata  witli  a  normal  maxiiiniiu 
of  about  21°-2.'i°  to  become  capable  of  livinp  at  70°    (see  Davenport). 

in  "Biochemistry  of  Muscle   and   Nerve,"   rhila..    1004. 

10  Quoted  by  irallilturlon. 


CAUSES  OF  NECROSIS  373 

may  be  more  susceptible  to  heat  than  under  other  conditions.  Just 
how  coagrulation  of  cell  o-lobulins  can  determine  the  deatli  of  a  cell 
is  difficult  to  understand,  unless  the  physical  conditions  of  the  cell 
are  jjreatly  altered  thereby.  Ordinarily  we  have  in  the  cell  an  equi- 
librium between  colloids  in  solution  and  colloids  in  the  solid  or  gel 
state;  if  the  colloids  are  rendered  insoluble  by  heat,  or  by  any  other 
cause,  so  that  this  equilibrium  is  destroyed,  serious  alterations  in  the 
mechanism  of  all  metabolism  must  result  (Mathews).  Other  chem- 
ical reactions  will  also  have  their  point  of  equilibrium  altered  by 
clian<>es  in  temperature,  and  such  .alterations  miglit  well  have  disas- 
trous results. 

Different  tissues  show  unequal  susceptibility  to  heat.  Werhov- 
sky  ^^  found  the  blood  most  affected  by  raising  the  temperature  of 
living  animals,  next  the  liver,  kidneys,  and  myocardium  in  order, 
the  other  tissues  being  little  or  not  at  all  structurally  injured. 

Cold  is  well  withstood  by  unicellular  forms,  and  relatively  poorly 
by  more  complex  organisms,  particularly  by  those  with  a  highly  de- 
veloped circulatory  system ;  this  is  because  individual  cells  are  not 
greatly  affected  by  freezing,  whereas  the  circulatory  channels  are 
readily  blocked  by  this  cause.  Bacterial  cells  are  not  killed  by  ex- 
posure for  long  periods  to  the  temperature  of  liquid  air  ^^  ( — 190°). 
Reduction  of  the  temperature  of  plant  cells  to  — 13°  may  result  in 
a  granular  transformation  of  the  cytoplasm,  often  with  rather  seri- 
ous structural  alterations.  Cytoplasm  seems  to  be  more  affected  than 
the  nucleus,  for  mitosis  may  occur  slowly  in  plant  cells  at  — 8°, 
and  Uschinsky  ^^  noted  that  in  animal  tissues  the  nuclei  were  less  af- 
fected by  cold  than  the  cytoplasm.  Blood  seems  little  affected  by 
freezing  temperature,  for  du  Cornu  found  that  dog's  blood  kept  on 
ice  for  five  to  ten  days  could  be  employed  for  transfusion  without 
causing  hemoglobinuria.  Grawitz  saw  motion  persist  in  human  cili- 
ated epithelium  kept  for  seven  to  nine  days  on  ice.  Ciliated  epi- 
thelium from  the  mouth  of  the  frog  may  survive  cooling  to  — 90°, 
and  frog  eggs  are  not  killed  by  — 60°.  In  many  cells,  however,  the 
physical  changes  produced  by  freezing,  and  also  bj^  the  subsequent 
thawing,  are  sufficient  to  render  them  incapable  of  further  exist- 
ence.-'' Cells  devoid  of  or  poor  in  water  cannot  be  killed  by  freez- 
ing, hence  it  is  probable  that  the  currents  set  up  about  the  crystals 
of  ice  in  thawing,  as  well  as  the  rapid  contraction  and  expansion 
under  the  influence  of  the  cold  and  the  ice  formation,  are  the  cause 
of  the  effects  of  freezing,  which,  therefore,  are  not  dependent  upon 
chemical,  but  upon  physical,  alterations. 

In  the  case  of  warm-blooded  animals,  the  gangrene  following  freez- 

i7Ziegler's  Beitr.,  1895   (18),  72. 
isMacFadven,  Lancet.  1900   (i),  849. 
i9Ziegler's  Beitr.,  1893   (12),  115. 

20  In  plant  cells  it  is  the  freezing  and  not  the  thawing  that  causes  the  harm 
(Maximow,  Berichte  Deut.  Bot.  Gesell.,  1912    (30),  504). 


374  h'i:Tii'(Kih'i:ssni-:  ciiAyaEH 

ing  depends  not  so  mueli  u])()n  the  freezing  of  the  cells  themselves  as 
upon  the  formation  of  hyalin  thrombi  in  the  injured  vessels  (v. 
Recklinghausen,  Hodara).-^  Kriege --  found  that  if  the  freezing  is 
transitory,  the  thrombi  may  again  disappear;  if  over  two  hours  in 
duration,  tliey  are  persistent.  Rischpler,-'  however,  considers  that 
cell  death  is  due  primaril}-  to  the  eft'ect  of  the  cold  upon  the  cells.  On 
tlie  other  hand,  Stechelmacher  -^^  found  that  freezing  of  liver  tissue 
produced  the  same  changes  as  ligation  of  the  hepatic  artery,  i.  e.,  in- 
creased permeability  of  the  cell  wall  followed  by  similar  changes  in 
the  nucleus,  suggesting  that  the  changes  produced  by  freezing  depend 
on  the  vascular  changes. 

Light.-"' — Light  may  atfect  tissues  seriously,  apart  from  the  efifects 
of  accompanying  heat,  although  the  experiments  of  xVron -^  indicate 
that  insolation  does  not  depend  on  the  light  rays,  but  solely  on  the 
heat.  Tn  the  treatment  of  lupus  by  the  Finsen  method  with  concen- 
trated light  rays,  the  action  is  largely  a  stimulating  one,  but  associ- 
ated with  or  subsequent  to  a  certain  degree  of  cell  injury.  Ogneff -'' 
found  that  moderate  action  of  electric  light,  rich  in  violet  and  ultra- 
violet rays,  causes  mitotic  cell  division :  if  the  action  is  stronger,  the 
cells  undergo  amitotic  division  and  then  become  necrotic.  Blue  rays 
have  but  slight  cytotoxic  action,  and  rays  further  towards  the  red  end 
of  the  spectrum  are  without  demonstrable  effect.  Light  l)aths  are  said 
by  Oerum  -•'  to  increase  greatly  the  quantity  of  corpuscles  and  hemo- 
globin, while  residence  in  the  dark  reduces  these  elements.  The  de- 
struction of  bacteria  by  light  is  a  well-known  phenomenon,-"  but  it 
has  been  suggested  that  their  destruction  depends  rather  upon  the 
action  of  substances  produced  in  the  culture-medium  under  the  influ- 
ence of  light  than  upon  the  effect  of  the  light  ujion  the  bacterial  cells 
themselves.  In  view  of  the  fact  that  enzymes  and  antibodies  in  solu- 
tion are  quite  readily  weakened  or  destroyed  by  the  action  of  light,  it 
is  possible  that  intracellular  enzymes  may  be  similarly  destroyed  by 
light,  with  resulting  cell  death.  However,  in  the  case  of  bacteria,  at 
least,  the  effects  of  light  seem  to  depend  upon  oxidation  processes,  for 
in  the  absence  of  oxygen,  bacteria  are  not  seriously  injured  by  light, 
and  D'Arcy  and  TIardy -*  fouiul  that  "active  oxygen"  is  formed  by 
the  same  ])()rtion  of  tlie  sjiectrum  that   is  most  active  in  destroying 

21  Miinch.  med.  Woch.,  1896   (43),  341. 

22Virc'how's  Arch.,  1880    (11(5),  f>4. 

23Ziefrl('r's  Beitr.,  1900   (2S),  .541. 

23aBoitr.  path.  Anat.,  1913   (57),  314. 

23bT{ovi(nv  l)y  Bering',  Er<ieb.  alljr.  Pathol..  1914,  Aht.  1.  (17).  790.  Soo  dis- 
cussion of  the  principles  of  the  action  of  lij^ht  on  tissues  h\'  Hox  if,  Anicr.  .lour. 
Tro])ical   Dis.,  191. I    (2).  500. 

2t  I'liilippine  .Tour.  Sci.,  B,  1911    (G).  101. 

2r.  I'lliijrer's  Arcli.,  1890   (03),  209. 

2<i  Plliifrer's  Arch..   190(1    (114),    1. 

27  Literature  fjrivcn  by  WiesiuT,  .\rch.  f.   llvg..  1907    Mil).   1. 

2x.l<)iir.  of   I'liysiol..  'lS!l.->    (17).   390. 


CAUSKH  OF  SECR08L-!  375 

Lacteria.-"  Li<iht  may  aiso  alter  tlie  soluliility  of  cell  jiroteiiis,  cspe- 
oiallj'  in  the  ]H"eseiK'e  of  various  organic;  and  iiior^nuiic  substances  that 
act  as  sensitizers,  such  as  silicates,  sug-ar,  lactic  acid  or  urea.-'"'  In 
this  may  lie  the  cause  of  cataract,  especially  diabetic  cataract. 

The  general  effect  of  "light  acting  on  oi-ganic  substances  present  in 
plant  and  animal  cells,  is  to  })roduce  from  earbonyl-containing  materi- 
als aldehyde  or  ketone  compounds,  whose  reactivity  and  availability 
for  important  synthetic  changes  are  conspicuous  (Xeuberg).^'' 
Whether  oxidative  processes  are  the  cause  of  death  in  animal  cells  is 
not  known,  but  we  are  familiar  with  many  chemical  reactions  of  vari- 
ous sorts  that  are  initiated  or  checked  by  the  action  of  light. ^^  Thus, 
bilirubin  is  oxidized  into  biliverdin,  when  acted  upon  by  sunlight, 
even  when  not  in  contact  with  air;  many  vegetable  oils  are  oxidized 
by  sunlight,  and  it  is  probable  that  the  oxidizing  action  of  light  upon 
organic  compounds  is  of  wide-spread  occurrence.  It  is,  therefore, 
quite  possible  that  such  oxidative  changes  may  be  the  cause  of  necrosis 
produced  by  the  action  of  light  rays,  especially  as  Bering  ^-  has  found 
that  chemically  active  light  rays  have  a  direct  action  on  oxidizing 
enzymes. 

It  is  very  probable  that  not  all  of  the  effects  of  exposure  to  the  sun 
depend  upon  the  heat  rays,  for  there  is  evidence  that  the  light  rays 
may  also  produce  effects.  This  is  definitely  true  in  the  case  of  indi- 
viduals or  animals  with  certain  pigments  in  their  blood,  notably 
liematoporphyrin  (q.  i\).  In  them,  not  only  may  skin  eruptions  re- 
sult from  relatively  small  exposure  to  light,  but  mice  may  be  so  sen- 
sitized that  a  few  moments  of  exposure  to  light  is  fatal. ^-'^  Artificial 
fluorescent  substances,  such  as  eosin,  also  sensitize  tissues  and  proteins 
to  light."-''  Normal  blood  absorbs  light  rays  in  large  amounts,  as 
Finsen  showed,  and  it  is  quite  possible  that  changes  in  the  ehemistrv 
of  the  blood  result  from  the  light  rays.  Exposure  to  the  sun  may 
cause  a  general  leucocytosis  with  relative  lymphocytosis.^-*^ 

According  to  Hertel  ^^  the  idtravlolet  rays  cause  oxygen  to  split  off 
the  easily  oxidizable  compounds  of  protoplasm,  and  Bovie  ^*  found 
that  they  coagulate  proteins ;  they  also  have  a  destructive  effect  on 
enzymes  -''  and  hormones.^^''  Toxins,  like  enzymes,  are  reduced  in 
activity  by  ultraviolet  rays.^*" 

29  See  also  Agulhon,  who  found  that  ultraviolet  ravs  may  attack  enzymes  to 
some  extent  in  the  absence  of  oxygen    (Ann.  Inst.  Pasteur..  1912    (20),  8S). 

29aSchanz,  Biochem.  Zeit.,  1915  (71),  406:  Burge.  Amer.  .Jour.  Phvsiol..  I'.tlti 
(39),  3.35. 

30  Biochem.  Jour.,  190S   (13).  305. 

31  See  Davenport.  "Experimental  ^forpliologv,"   1897,  p.   102. 
32Miinch.  med.  Woch..  1912    (59).  2795. 

32aHausmann,  Biochem.  Zeit..   1914    (67).  309. 

32b  Pincussohn,  Deut.  med.  Woch.,   1913    (44),  2143. 

32c  Aschenheim.  Zeit.  Kinderheilk.,  1913    (9),  87. 

33  Zeit.  AuGfenlieilk.,  1911    (26),  393. 

34  Science,  1913   (37),  24:  see  also  Burge.  Amer.  Jour.  Plivsiol.,  1910   (39).  335. 
34ii  Burge  et  a/.,  Ainer.  Jour.  Physiol.,  1916    (40).  426. 

34bHartoch  ef  nl..  Zeit.  Immunit'iit.,  1914   (21),  643. 


376  RETROGRESSIVE    CHANGES 

X=rays  ^"  stimulate  cell  growth  when  applied  in  small  amounts,^^" 
hut  larger  amounts  produce  necrosis,  which  is  peculiar  in  that  an  in- 
tem'al  of  several  days,  or  even  weeks,  may  elapse  after  the  exposure 
before  the  necrosis  manifests  itself.  Ellis  ^^^  considers  that  the  amount 
of  necrosis  is  out  of  proportion  to  the  changes  in  the  vessels,  which 
some  have  believed  to  be  the  cause  of  a--ray  gangrene,  and  therefore 
that  the  cells  must  be  directly  injured,^"  a  view  supported  by  Cascr 
mir's^^  experiments  with  plant  cells.  The  extensive  studies  of  the 
Hertwigs  show  that  the  chromatin  is  chiefly  affected,  which  presum- 
ably explains  the  fact  that  immature  cells,  and  cells  in  active  division, 
are  more  sensitive  to  ic-rays  than  adult  cells,  and  that  monstrosities 
develop  from  eggs  exposed  to  radiant  energy.  As  far  as  histological 
changes  show,  hard  rays  produce  less  but  quite  the  same  changes  as 
soft  rays.  That  .r-rays  have  a  marked  effect  on  metabolism  has  been 
abundantly  established.^*  According  to  ]\Iusser  and  Edsall,^^  the  ef- 
fect of  ar-rays  upon  metabolism  is  unequalled  by  any  other  therapeu- 
tic agent,  and  is  manifested  by  excessive  elimination  of  the  products 
of  protein  destruction,  which  arise  particularly  from  the  lymphatic 
structures."  These  changes  have  been  studied,  therefore,  particu- 
larly in  connection  with  the  treatment  of  leukemia  (q.  v.).  In  con- 
sequence of  the  injuiy  to  the  blood-forming  tissues,  resistance  to  bac- 
teria is  decreased  (Lawen).*^  The  renal  epithelium  seems  also  to 
suffer  injury  in  some  cases.*- 

Badiiun,  which  shares  with  x-rays  the  power  of  causing  tissue 
necrosis,  does  not  have  a  similar  effect  upon  the  blood,  nor  do  the 
ultra-violet  rays  (Linser  and  Helber).^^  In  general,  however,  radium 
has  much  the  same  effect  on  tissues  as  .r-rays,**  but  seems  rather  to 

35  Full  review  by  Cohvell  and  Russ,  "Radium,  X-Rays  and  tlio  Living;  Cell," 
London,  1915.     Also  see  Richards.  Science,  1915    (42),  287. 

3oaSee  Schwarz,  Miinch.  med.  Woch.,  1913    (60),  21G5. 
3-.b  Amer.  -Tour.  :\red.  Sci.,  190.3    (125).  85. 

36  Allen  (Jour.  :Med.  Research,  1903  (9),  402)  states  that  i)rotnzoa  and  vinegar 
eels  are  killed  by  long  exposure  to  ir-rays,  wheieas  plants  are  decidedly  stimulated 
in  their  growth. 

37  Med.-Xaturw.  Arch.,  1910   (2).  423;  r^sum^  on  iP-rays. 

■'■^  See  Harvey  (Jour.  Path,  and  Bact.,  1908  (12),  548)',  concerning  the  effects  of 
a;-rays. 

3o'Univ.  Penn.  Med.  Bull.,  1905  (IS),  174:  also  Edsall  and  Pemberton.  Amer. 
Jour.  Med.  Sci.,  1907    (133),  426. 

•«o  A  peculiar  selective  action  for  the  generative  cells  is  also  shown  by  .r-rays, 
which  cause  marked  atrophy  of  the  ovaries  and  testicles.  In  the  latter  it  aflVcts 
chiefly  the  gerniinative  cells,  spari'i'  Mic  cells  of  Levdig.  (See  Albers-Schonbcrg, 
Miinch.  med.  Woch.,  1903  (50),  185'):  Frieben,  ihid..  1903  (50),  2295;  Spccht. 
Arch.  f.  Gvn..  1906  (78),  458;  Tlialer.  Deut.  Zeit.  f.  Chir.,  1905  (79),  576;  Rcif- 
ferscheid,  Zeit.  f.  Cvn.,  1910   (34),  593. 

■ti  Mitt.  Grenz.  Med.  u.  Chir.,  1908    (19),   141. 

42  See  Schulz  and  Hoffman,  Deut.  Zeit.  f.  Chir.,  1905  (79),  350:  Warthin,  Amer. 
Jour.  :Med.  Sci..  1907    (133),  736. 

■•3  Deut.  Arch.  klin.  Med.,   1905    (S3),  479. 

44  Review  bv  Cuvot,  Cent.  allg.  Path.,  1909  (20),  243;  also  see  :Mills.  Lancet. 
1910   (179),  462;  Ricliards,  Science,  1915   (42),  287. 


CAUSES  OF  NECROSIS  377 

stimulate  the  action  of  most  enzymes;*'^  autolysis,  however,  is  iiol  in- 
creased (Brown).*"  In  proper  amounts  radium  stimulates  })lant 
metabolism  (Gager).  Thorium-a;  also  attacks  specifically  the  leuco- 
cytes,*®" so  that  by  proper  dosage  an  animal  may  be  made  practically 
leucocyte-free,''"''  which  has  been  used  for  experimental  studies  on  the 
functions  of  the  leucocytes. 

The  lonpr-continued  action  of  a:-rays  upon  the  skin  has,  in  many 
cases,  led  to  the  formation  of  cancer,  apparently  because  the  pro- 
liferation stimulated  by  the  rays  progresses  until  it  exceeds  normal 
bounds. ^^  Likewise  leukemia  has  been  observed  several  times  in 
roentgenologists,  presumably  produced  in  the  same  way.**"  Radium 
also  causes  severe  skin  lesions  and  a  general  lymphocytosis  in  those 
exposed  to  it  for  long  periods.**'' 

As  the  metabolic  changes  produced  by  a'-rays  indicate  an  extremely 
high  rate  of  autolysis,  one  may  ascribe  the  effects  either  to  a  stimulat- 
ing effect  of  x-rays  upon  autolytic  enzymes,  or  as  Neuberg*"  does,  to 
an  inhibitive  action  of  .r-rays  and  radium  rays  upon  the  other  intra- 
cellular enzymes  wnthout  a  corresponding  deleterious  effect  upon  the 
autolytic  enzymes."'"  This  hypothesis  agrees  with  the  facts  at  hand, 
but  more  details  concerning  the  effects  of  these  rays  upon  various 
enzymes  are  needed.  The  long  latent  period  before  the  appearance 
of  necrosis  after  exposure  to  rr-rays  is  difficult  to  explain,  and  agrees 
rather  wath  the  hypothesis  of  slow  proliferative  and  obstructive 
changes  in  the  blood-vessels. 

Electricity. — The  effects  of  the  electric  current  upon  cells  are  de- 
scribed by  Davenport  as  follows:  A  weak  constant  current  causes 
a  centripetal  flowing  of  the  protoplasm  (in  Actinosphaerium)  ;  if  the 
current  is  increased  or  long  continued,  the  cytoplasm  of  the  pseudo- 
podia  becomes  varicose,  and  droplets  are  formed  which  soon  burst, 
causing  a  collapse  of  the  protoplasmic  framework.  Finally,  the  pro- 
toplasm on  the  anode  side  begins  to  disintegrate,  and  the  loose 
particles  move  toward  the  positive  electrode;  eventually  the  cell  struc- 
ture may  be  entirely  destroyed.  A  similar  disintegration  of  the 
anode  side  of  ameba  has  been  observed  by  McClendon,^^   which  he 

45  Loewenthal,  Berl.  klin.  Woch.,  1010  (47),  287:  Kionka,  Med.  Klinik,  1911 
(7),  685.     Denied  bv  Gudzent,  Zeit.  Strahlontlier.,  1014    (4),  666. 

46  T.  R.  Brown,  Arch.  Int.  Med.,  1012   (10),  405. 

46a  See  Plesch  et  al.,  Zeit.  exp.  Path.,  1012   (12),  Xo.  1. 

46b  There  is  no  increase  in  antitrypsin  from  this  leucocyte  destruction  (Rose- 
now,  Zeit.  exp.  Med.,  1014   (3),  377)". 

47  See  review  bv  Wyss.  Beitr.  z.  klin.  Chir..  1006  (40),  185;  Porter  and  Wol- 
bach,  Jour.  Med.  Res.',  lOW)   (21),  357. 

48  See  Jagic  and  Schwarz,  Berl.  klin.  Woch..  1011    (48),  1220. 
48a  See  Ordway,  Jour.  Anier.  Med.  Assoc,  1016  (66),  1. 

49  Zeit.  f.  Krebsforschung,  1004  (2),  171;  also  Meyer  and  Berins;,  Fortschr. 
Roentg:enstrahlen,  1011    (17),  33;  Richards,  Amer.  Jour.  Physiol.,  1014   (36).  400. 

50  Some  authors  have  believed  certain  of  tlie  effects  of  a^-rays  to  be  produced 
by  choline  liberated  throufrh  the  decomposition  of  lecithin.  (See  Benjamin  and 
Reuss,  Miinch.  med.  Woch..  1006   (53),  1860.) 

BiPfliiger's  Arch.,  1011    (140),  271. 


378  RETROGREf^SIVE    CH.WGES 

attributes  to  anions  which  cannot  pass  through  the  cell  wall,  and 
therefore  accumulate  on  that  side  of  the  organism.  If  an  alter- 
nating current  is  used,  both  anode  and  cathode  sides  of  the  cell  are 
affected.  In  moving  organisms  electric  currents  determine  direction 
of  motion,  even  certain  vertebrates  (tadpoles,  fish)  being  made  to 
orient  themselves  according  to  the  current.  The  nucleus  seems  to  be 
more  susceptible  to  harm  by  electric  currents  than  the  cytoplasm 
(Pfeffer),^'-  and  there  seems  to  be  no  oxidation-process  involved  in 
cell  destruction  by  electricity  (as  is  the  case  with  light  rays),  for  the 
effects  are  much  the  same  in  the  absence  of  oxygen  (Klemm). 
Schmaus  and  Albrecht  state  that  the  effect  of  electricity  upon  proto- 
plasm depends  upon  a  loosening  of  the  cohesion  and  a  solution  of  the 
constituents  of  the  cell  (vacuolization),  which  last  is,  perhaps,  due 
to  direct  chemical  alterations.  It  may  be  suggested  that  the  electric 
current  causes  a  migration  of  ions  toward  one  or  the  other  pole 
of  the  cell,  in  this  way  separating  the  movable  inorganic  ions  of  the 
ion-protein  compounds  of  the  cell  from  the  immobile  colloidal  pro- 
teins, with  consequent  serious  alterations  in  the  chemistry  of  the  cell. 
Zeit  ^^  found  that  continuous  currents  kill  bacteria  through  the  pro- 
duction of  antiseptic  substances  in  the  culture-medium,  but  do  not 
harm  them  directly. 

Jellinek  •'*  has  studied  extensively  the  cause  of  death  after  severe 
electric  shocks,  and  finds  that  there  are  produced  intracerebral  hemor- 
rhages and  degeneration  of  the  nerve-cells,  which  are  sufficient  to 
explain  the  death  of  the  individual  without  having  recourse  to  the 
more  indefinite  idea  of  "shock."  Cunningham  ^^  considers  fibrillary 
contraction  of  the  heart  as  the  cause  of  death.^*'  Spitzka  and 
Radasch  "  find  changes  in  the  brains  of  electrocuted  criminals,  which 
indicate  a  sudden  liberation  of  gas  about  the  blood  vessels,  along  which 
the  current  passes.  The  amperage  seems  to  be  far  more  important  in 
determining  the  effect  of  a  current  than  the  voltage  or  wattage."''-'^ 

Chemicals  cause  cell  death  whenever  they  are  of  such  a  nature  as 
either  to  coagulate  the  cell  proteins  or  to  destroy  its  enzymes.  The 
action  of  such  substances  as  sulphuric  acid,  strong  caustics,  etc., 
hardly  calls  for  ex])lanation.  Phenol  (carbolic  acid)  may  cause  ne- 
crosis and  gangrene  even  when  in  very  dilute  solution ;  this  appears 
to  be  due  more  to  the  production  of  hyaline  thrombi  of  agglutinated 
red  corpuscles  in  the  capillaries  than  to  direct  action  upon  the  cells. 
In  some  unpublished  experiments  on  the  subject  of  "carbolic  acid 
gangrene,"   1   found  this  action  of  phenol  very  striking  when  dilute 

52  Literature  given  bv  Davenport.  "Experimental  Morphology." 
•"■3  .Tour.  Amer.  Mod.  Assoc.  1001    (.37),  1432,  literature. 
S4  Vircliow's  Arcli.,  1002    (170),  oO;  Lancet,  1003    (i),  357. 
n.iXew  York  Med.  .lour.,   ISOO    (70),  'jSl. 

•'^»  T'^lll  discussion  bv  .TelliJTe  in  Peterson  and  irainos"  "Loiral  ^lediciiio  and 
Toxicologv,"  1003    (1)"24.'5. 

57  Amer.  .Tour.  Med.  Sci.,  1012   (144),  341. 

57a  Jellinek,  Wien.  klin.  Woeli.,  1013   (20),  170;{. 


CAUSES  OF  NECROSIS  379 

solutions  were  plaeed  on  the  web  of  a  frog's  foot,  under  the  micro- 
scope; as  soon  as  tiie  solution  penetrated  to  a  capillary,  stasis  with 
fusion  of  the  corpuscles  occurred  in  a  very  few  seconds.  Similar 
results  have  been  obtained  by  Rosenberger.'*'*  Some  poisons  seem  to 
cause  necrosis  without  destroyiiiu'  the  autolytic  enzymes,  in  wliicli 
ease  the  cells  are  rapidly  digested ;  at  least,  such  a  hypothesis  seems 
to  explain  best  the  changes  seen  in  the  liver  in  chloroform  poisoning, 
acute  yellow  atrophy,  eclampsia,  etc^**  Not  all  poisons,  by  any 
means,  cause  cell  death — tetanus  toxin,  morphine,  and  other  alkaloids 
cause  death  of  the  individual  as  a  whole  without  usually  causing  pri- 
mary necrosis  of  any  of  the  cells.  Cell  death  does  not  necessarily  de- 
pend upon  destruction  of  all  the  cellular  enzymes,  as  has  been  pointed 
out  previously.  Thus,  bacteria  may  be  killed  by  many  chemicals 
which  seem  not  to  att'ect  their  autolytic  enzymes  seriously.  Any  con- 
siderable excess  of  either  H  or  OH  ions  is  incompatible  with  cell  life, 
and  it  is  possible  that  at  times  the  production  of  acids  within  a 
cell  may  be  sufficient  to  cause  death  °°^  (e.  g.,  in  the  kidney  in  acute 
nephritis  (]\I.  H.  Fischer),  or  in  the  muscle  in  waxy  degeneration 
(Wells) ■'"''■"'.  It  is  quite  probable  that  many  of  the  poisons  act  by 
interfering  with  the  oxidative  capacity  of  the  cells;  this  seems  almost 
certain  in  the  case  of  chloroform  necrosis,  and  even  bacterial  poisons 
(diphtheria  and  typhoid)  were  found  by  Pitini  *'°  to  decrease  the 
oxidizing  power  of  the  cells. 

The  term,  "protoplasmic  poison,"  has  been  variously  used  and  de- 
fined. Kunkel  says  that  a  protoplasmic  poison  "is  a  poison  which, 
without  producing  directly  evident  alterations,  harms  or  kills  all 
living  protoplasmic  structures."  HgCU  is  such  a  poison,  whereas 
HoSOj,  bromine,  and  similar  substances  that  destroy  all  life  through 
their  strong  chemical  action  are  not  included  in  this  category.  The 
protoplasmic  poisons  presumably  act  by  combining  with  one  or  more 
of  the  constituents  of  cell  protoplasm;  e.  g.,  HgCL  probably  combines 
with  the  proteins,  chloroform  with  the  cell  lipoids  (physicall}'?).  By 
means  of  his  special  technic  Barber  ^^  is  able  to  introduce  minute 
quantities  of  poisons  into  living  cells  and  observe  their  effect  on  the 
cytoplasm :  ITgCL,  is  thus  found  to  be  most  toxic,  while  ASoO.j  is 
relatively  inert.  Mathews "-  has  shown  that  the  toxicity  of  ions 
depends  on  the  ease  with  which  they  part  with  their  electrical  charges, 

ssVerh.  Phys.  Med.  Gesellsch.  z.  Wiirzburor,  inoo.  vol.  34. 

59  Wells,  Jour.  Amer.  Med.  Assoc.  1006   (46),  .341. 

59a  The  partial  protection  afforded  by  a  rich  carbohydrate  diet  against  the 
necrogenic  action  of  chloroform,  phosphorus  and  renal  poisons,  as  observed  by 
Opie  and  Alford  (Jour.  Exp.  ^Med.,  1915  (21),  1),  may  depend  on  the  anti- 
ketogenic effect  of  carbolivdrates. 

59b  Jour.  Exp.  Med.,  inoO    (11),   1. 

eoBiochem.  Zeit.,   1910    (25),  257. 

61  Jour.  Infect.  Dis.,  1911    (9),  117. 

62  Amer.  Jour.  Physiol.,  1904  (10),  290;  Nicholl.  Jour.  Biol.  Cliem.,  1909  (5), 
453. 


380  RETROGRESSIVE    CHANGES 

and  the  toxicity  of  a  salt  is  a  function  of  the  sum  of  the  toxicity 
of  the  ions;  hence  the  toxicity  of  a  salt  is  in  inverse  proportion  to 
its  decomposition  tension.  Kunkel  suggests  that  oxalic  acid  and 
fluorides  are  poisons  because  they  combine  the  cell  calcium,  and 
barium  salts  may  be  poisonous  because  they  precipitate  the  SO4  ions. 
We  can  readily  imagine  that  the  combining  of  even  one  of  the  essential 
constituents  of  the  cell  may  so  upset  the  normal  chemical  processes 
that  the  cell  can  no  longer  take  up  substances  to  repair  its  waste,  and 
hence  necrosis  ensues.^^ 

Physical  agents  may  cause  necrosis,  usually  in  ways  too  obvious 
to  require  explanation.  With  most  cells,  large  portions  of  the  cyto- 
plasm can  be  destroyed  without  serious  results,  for  so  long  as  the 
nucleus  is  intact  the  cytoplasm  can  be  reconstructed.  The  fact  that 
necrosis  frequently  follows  relatively  slight  injuries  of  the  nucleus 
is  perhaps  best  explained  by  considering  that  injury  to  the  nuclear 
membrane  modifies  the  permeability  of  the  nucleus  for  substances  in 
solution,  which  might  readily  affect  its  metabolic  activities  to  a  serious 
degree.  It  is  possible,  also,  that  solvents  of  lipoids,  such  as  chloro- 
forai,  etc.,  produce  much  of  their  deleterious  effects  by  modifying 
the  permeability  of  the  cell,  if  the  semipermeability  of  cell  mem- 
branes depends  largely  upon  the  lipoids  they  contain."* 

Physical  injury  of  even  slight  degree  may  bring  on  severe  alterations 
in  cells,  however,  and  indeed  may  cause  severe  chemical  alterations. 
We  know  that  many  chemical  reactions  can  be  brought  about  by  slight 
mechanical  disturbances,  e.  g.,  the  explosion  of  fulminate,  nitrogen 
iodide,  etc.,  and  it  is  quite  possible  that  mechanical  disturbances  can, 
likewise,  cause  chemical  changes  in  the  protoplasm.  ]\Ian3^  lower 
animals  devoid  of  a  nervous  system  respond  to  mechanical  stimuli 
by  chemical  activity ;  e.  g.,  the  production  of  phosphorescence  by 
marine  organisms  when  agitated  by  an  oar,  etc.  Possibh%  the  secre- 
tion of  thrombokinase  by  the  leucocytes,  which  occurs  whenever  they 
come  in  contact  with  a  foreign  body,  is  an  example  of  a  similar  re- 
action to  a  mechanical  stimulus.  Even  in  urticaria  factitia  the  sim- 
ple mechanical  irritation  which  suffices  to  produce  the  wheals  is 
followed  very  quickly  by  extensive  nuclear  fragmentation,^^  but  it  may 
be  that  unknown  poisons  are  present  in  the  hypersensitive  skin  and 
cause  the  karyorrhexis,  and  not  the  trauma  alone.  We  have  no  good 
evidence  that  mere  contact  with  a  chemically  inert  foreign  body  uiuic- 
com]ianied  by  cellular  injury,  can  cause  death  of  tissue-cells."" 

Extreme  changes  in  osmotic  pressure  may  lead  to  cell  death,  either 

«3  It  is  hardly  profitablo  here  to  go  further  into  tlie  theories  of  the  aetion  of 
poisons,  which  are  generally  extensively  considered  in  the  treatises  on  toxicology 
and  pharmacology    (also  by  Davenport,  loe.  cit.) . 

"4  See   I'ascncci*.  Tlofmeister's  IJeitriigc,   190")    (0),  ,'552. 

"r- Gilchrist,  Bull.  Johns  Hopkins  Tlosp.,  1008    (19),  49. 

•■■"Meltzer  (Zeit.  f.  Riol.,  1894  (.30),  4(14)  has  shown  tliat  bacteria  may  be 
killed  by   violent    agitation,   whicli   causes  disintegration  of  tlie  cells. 


VAIUETlEii  OF  \ECltOSIS  381 

b}^  causing  structural  alteration  in  the  cell  (c.  <j.,  the  bursting;  of 
plant-cells  in  water),  or  concentration  of  the  electrolytes  may  become 
so  great  that  the  colloids  are  thrown  out  of  solution,  as  in  the  ordi- 
nary salting-out  processes  of  the  laboratory.  It  is  doubtful,  however, 
if  osmotic  changes  per  se  ever  become  so  abnormal  within  the  animal 
body  (except  in  experimental  conditions)  as  of  themselves  to  cause 
cell  necrosis. 

VARIETIES  OF  NECROSIS 

Coagitlation  Necrosis.''" — This  name  is  applied  to  necrotic  areas 
that  are  lirni,  dry,  usually  pale  yellowish  in  color,  and  observed  prin- 
cipally in  areas  of  total  anemia  or  tuberculosis.  The  question  has 
been  long  disputed  as  to  whether  a  true  coagulation  occurs  in  such 
tissues  or  not.  Necrosis  produced  by  heat,  carbolic  acid,  corrosive- 
sublimate,  etc.,  is  naturally  a  coagulation  necrosis,  the  cells  of  the 
att'ected  area  having  undergone  true  coagulation ;  i.  e.,  the  conversion  , 
of  their  soluble  colloids  {sols)  into  the  insoluble  ''pectous"  modifica- 
tion. Whether  the  same  change  occurs  in  areas  of  anemic  necrosis 
is  not  so  well  established.  If  the  part  contains  a  fair  amount  of 
plasma  the  liberation  of  the  tissue  coagulins  from  the  dead  cells  will 
cause  a  conversion  of  the  fibrinogen  into  fibrin — this  can  usually  be 
demonstrated  microscopically,  but  the  presence  of  fibrin  is  not  con- 
stant, and  its  quantity  is  usually  insufficient  to  explain  satisfactorily 
the  condition  of  coagulation  necrosis  in  infarcts,  etc.,  as  Weigert 
maintained."^  Sclimaus  and  Albrecht  believe  that  a  true  coagula- 
tion of  the  cell  proteins  does  occur  in  anemic  infarcts,  etc.,  for  they 
found  that  the  cells  of  kidneys  with  ligated  vessels  contain  at  first 
granules  soluble  in  water  and  salt  solution ;  after  forty-eight  hours 
the  granules  cannot  be  dissolved  in  these  solvents  or  in  weak  acetic 
acid,  but  are  soluble  in  2  per  cent.  KOH ;  after  five  to  six  days  the 
granules  are  insoluble  even  in  KOH.  Beyond  these  experiments,  we 
seem  to  have  no  proof  of  the  occurrence  of  intracellular  coagulation 
within  areas  of  coagulation  necrosis  due  to  anemia :  exact  chemical 
studies  on  this  point  are  much  needed.  Since  tissue-cells  contain 
coagulins  for  fibrinogen,  it  is  possible  that  they  also  contain  coag- 
ulins for  cell-proteins,  but  this  remains  to  be  established.  Bacteria 
produce  substances  coagulating  milk  and  fibrinogen.  Bergey  '''*  calls 
attention  to  the  coagulation  of  serum  by  enzymes  and  acids  produced 

67  Literature  J)y  Jores,  Ergebnisse  der  Pathol.,  180S    (5),  16. 

68  Weigert  believed  that  the  dead  area  becomes  permeated  by  plasma  eontainiiig 
fibrinogen,  which  is  coagulated  in  and  between  the  cells.  lie  put  much  weiglit 
on  an  increase  in  size  of  the  necrotic  area,  wliich  is  by  no  means  constant,  as  he 
i-itimated;  necrotic  areas  are  inelastic,  and  when  death  occurs  tliey  do  not 
shrink  with  the  fall  of  lilood  juessure  as  the  surrounding  tissues  do,  and  lience 
they  may  appear  to  project  from  the  surface  of  the  dead  organ  Mlien  tiiey  did 
not  do  so  dviring  life.  According  to  Moos  (Virchow's  Archiv.,  l!)On  (10.5) ,  27.'}) 
the  plasma  does  not  permeate  infarcted  areas  to  the  e.xtent  that  ^Yeigert  assumed. 

fisJour.  Amer.  Med.  Assoc.,  15107    (40),  G80 


382  RETROGRESSIVE    CHANGES 

by  bacteria,  and  Ruppel  '"  found  that  the  tubercle  bacillus  produces 
substances  precipitating-  proteins ;  hence  coagulation  necrosis  in  bac- 
terial infections  may  be  brought  about  in  this  way,  and  SchmoU  "^ 
has  shown  that  the  necrosis  occurring  in  tubercles  is  associated  with 
an  almost  complete  coagulation  of  the  cell-proteins. 

Necrosis  associated  with  infiannnatory  exudation  is,  of  course,  ac- 
companied by  coagulation  of  the  fibrinogen  of  the  exudate  (e.  g., 
diphtheria)  ;  this  type  of  coagulation  necrosis  is  chemically  a  simple 
Mbrin-formation  and  readily  understood.  The  peculiar  hyaline  de- 
generations of  parenchymatous  cells  {e.  g.,  Zenker's  degeneration 
of  muscles)  are  often  included  under  this  class,  but  it  would  seem 
more  probable  that  the  processes  consist  rather  of  the  fusion  of  the 
structural  elements  of  the  cell  into  a  homogeneous  substance  than  a 
true  coagulation.  When  necrosis  is  produced  by  chemical  means 
more  or  less  coagulation  of  some  of  the  soluble  proteins  probably 
takes  place ;  even  in  plant  cells  this  coagulation  of  dead  protoplasm 
is  described.'- 

Liquefaction  necrosis  occurs  particularly  in  the  central  nervous 
system,  where  the  cell  substance  seems  not  to  undergo  the  coagula- 
tive  changes  described  in  the  preceding  paragraphs.  Whether  this  is 
due  to  a  lack  of  tissue-coag-ulins  or  to  a  difference  in  cell  composition 
cannot  be  said,  but  the  large  proportion  of  lipoids  in  brain  tissue  is 
probably  an  important  factor.  Probably  "edema  ex  vacuo"  is  re- 
sponsible for  much  of  the  accumulation  of  fluid,  due  to  the  anatomical 
conditions  that  prevent  a  shrinking  or  collapse  of  the  tissues  to  fill 
in  the  gap,  and  the  lack  of  connective-tissue  formation.  Aseptic 
softening  in  general  may  be  safely  ascribed  to  digestion  of  proteins 
by  cellular  enzymes,  either  from  the  dead  cells  or  from  the  leuco- 
cytes. Suppuration  is  merely  a  form  of  liquefactive  necrosis,  in 
which  such  digestion  is  particularly  rapid  because  of  the  large  num- 
ber of  leucocytes  that  are  present.  Necrosis  of  the  gastric  mucosa  or 
of  the  pancreas  is  also  followed  by  rapid  liquefaction,  through  the 
action  of  the  digestive  enzymes  of  these  tissues.  When  necrosis  is 
accompanied  by  edema  (as  in  superficial  burns),  the  fluid  enters  the 
cells  in  large  amounts,  and  in  this  way  another  form  of  liquefaction 
necrosis  may  be  produced.  Bacterial  enzymes  may  be  a  factor  in  pro- 
ducing liquefaction  of  dead  tissues,  but  with  most  pathogenic  foinns 
there  is  little  proteolytic  activity. ^^ 

Caseation. — This  term  is  applied  to  a  form  of  coagulation  necrosis 
in  which  the  dead  tissue  has  an  appearance  quite  similar  to  that  of 
cheese.  If  we  bear  in  mind  the  fact  that  cheese  is  a  mixture  of 
coagulated  protein  and  finely  divided  fat,  and  that  in  caseation  we 

ToZeit.  phvsiol.  Cliem..   1898    (26),  218. 

71  Dent.  Arch.  klin.  Vvd.,  in04   (81),  lt!.S. 

■^  (laiaukov,  Zcit.  clioni.  Kolloide,   1!»1()    (fi),  ^(in ;    l.op.'sclikin.   Bor.  DcMit.  Bot. 
(Jesell.,  1012    CM)),  .')28. 
■    73  See  Bittrolff,  Beitr.  path.  Anat.,  101.')   (fiO),  X^7. 


\.\nn:T/i:s  of  xecrosis  383 

have  a  coagulation  of  tissue  proteins  associated  with  the  deposition 
of  considorahle  quantities  of  fat,  the  reason  for  the  gross  roseniljhince 
of  the  pro(hic't  of  tliis  form  of  necrosis  to  cheese  is  ai)i)arent. 
Schmoll  '*  has  analyzed  caseous  nuiterial,  and  found  it  almost  en- 
tirely free  from  soluble  proteins  or  proteoses.  The  protein  material 
is  almost  solely  coagulated  protein,  which  in  its  elementary  composi- 
tion is  related  to  the  simple  proteins  or  to  fibrin,  and  not  at  all  to 
the  luicleoproteins.  The  extremely  small  amount  of  phosphorus  pres- 
ent in  the  caseous  material  indicates  that  the  products  of  disintegra- 
tion of  the  cell  nuclei  must  diffuse  out  earh^  in  the  process.  Casea- 
tion is,  therefore,  characterized  by  a  coagulation  of  the  proteins  and 
a  dissolving  out  of  the  nuclear  components.  SchmoU  does  not  ex- 
plain the  cause  of  coagulation,  however.  It  may  be  that  it  is  the 
same  as  in  the  coagulation  of  anemic  infarcts  (since  tuberculous  areas 
are  decidedly  anemic),  or  possibly  the  tubercle  bacillus  produces  sub- 
stances coagulating  proteins,  as  Ruppel  states  is  the  property  of 
' '  tuberculosamin. ' '  Indeed,  Auclair  ' "'  claims  that  the  fatty  sub- 
stance that  can  be  extracted  from  tubercle  bacilli  by  chloroform  is 
the  cause  of  the  caseation.  Dead  tubercle  bacilli  do  not  produce  true 
caseation,  however,  according  to  Kelber ;  ''^  hence  the  substance  caus- 
ing the  necrosis  evidently  does  not  diffuse  readily  from  the  bodies  of 
the  bacilli. 

The  abundance  of  fat  in  caseous  material  is  very  striking.  Bos- 
sart  "  found  from  13.7  per  cent,  to  19.4  per  cent,  of  the  dry  sub- 
stance of  caseous  material  soluble  in  alcohol  and  ether.  In  the  scrap- 
ings from  tuberculous  bovine  glands  I  have  found  22.7-23.9  per  cent, 
of  the  organic  material  soluble  in  alcohol  and  ether.'*  Of  this  solu- 
ble material,  Bossart  found  25  to  33  per  cent,  of  cholesterol,  and 
Leber  '°  found  38.31  per  cent,  of  lecithin,  which  is  a  much  higher 
proportion  than  Bossart  detected.  Presumably  these  fatty  materials 
are  derived  chiefl.y  from  the  disintegrated  cells;  this  is  probably  true 
of  the  lecithin  and  cholesterol,  but  the  fact  that  in  histological  prepa- 
rations most  of  the  fat  is  found  about  the  periphery  of  the  caseous 
area,^'^  supports  the  belief  that  it  has  wandered  in  from  the  outside. ^^ 
A  certain  proportion  of  the  fat  is  possibly  derived  from  the  bodies 
of  the  tubercle  bacilli,  which  usually  contain  about  40  per  cent,  of 
fatty  matter;  but  it  has  not  been  determined  whether  the  fat  from 
this  origin  forms  an  appreciable  part  of  the  fatty  matter  of  caseous 
material. 

'•*Deut.  Arch.  klin.  :\red..  1004   (81),  163. 

"5  Arcli.  med.  expi^r..  ISitD,  p.  .SCi.S. 

78  Quoted  by  Diirck  and  Oheriidorfer,  Ergebnisse  der  Pathol.,  1800    (0),  288. 

77  Quoted  bv  Sclimoll,  Joe.  cif.'i 

78  Wells.  -Tour.  :\led.  Kesearch,  1006   (14),  401. 

79  Quoted  bv  Solinioll.74 

soSata,  Ziejiler's  Beitr.,  1000   (28).  461. 

81  Fischler  and  Gross  (Zieoler's  Beitr.,  1005  (7th  suppl.),  344)  could  find  no 
fatty  acids  in  caseous  areas  by  histological  metliods. 


384  RETROGRESSIVE    CHANGES 

Caseous  areas  persist  for  extremely  long  periods  of  time  without 
undergoing  absorption,  which  indicates  that  the  autolytic  enzymes 
are  destroyed  early  in  the  process,  presumably  b}^  the  toxins  of  the 
tubercle  bacillus;  corresponding  to  this  SchmoU  found  autolysis  very 
slight  indeed  in  caseous  areas,  and  even  wiieii  tlie  caseous  material 
breaks  down  to  form  a  "cold  abscess"  the  fluid  differs  from  true  pus 
in  containing  less  free  amino-acids,  e.  g.,  tyrosine  is  mission.*-  Be- 
cause of  a  lack  of  chemotactic  substances  no  leucocytes  enter  to  re- 
move the  dead  material,  in  consequence  of  wliich  caseous  material  gives 
no  evidence  of  containing  proteases,  according  to  the  Miiller-Joch- 
mann  ])late  method.  That  the  failure  of  absorption  is  not  due  to 
a  modification  of  the  proteins  into  an  indigestible  form  is  shown  by 
the  rapid  softening  of  caseous  areas  when,  through  mixed  infection, 
chemotactic  substances  are  once  developed  and  leucocytes  enter.  Job- 
ling  and  Petersen  ^-^  suggest  that  in  caseation  the  autolysis  is  inhibited 
by  the  soaps  of  fatty  acids,  which  are  abundant  in  caseous  areas  and 
have  a  marked  antitryptic  effect. 

FAT  NECROSIS  83 

Through  usage  this  term  has  come  to  indicate  a  specific  form  of 
necrosis  of  fat  tissue,  which  is  characterized  by  a  focal,  circum- 
scribed arrangement,  and  by  the  splitting  of  the  fat  in  the  necrotic 
area  into  fatty  acids  and  glycerol,  the  latter  disappearing,  the  former 
combining  with  bases  to  form  soaps.**  In  practically  all  cases  fat 
necrosis  is  produced  by  the  action  of  pancreatic  juice  upon  fat  tis- 
sue,*^ presumably  through  the  action  of  the  enzymes  it  contains,  and 
the  condition  can  be  produced  experimentally  by  any  procedure  that 
causes  escape  of  the  pancreatic  juice  from  its  natural  channels. 

Langerhans  *^  made  the  first  studies  of  the  nature  of  the  changes 
in  fat  necrosis,  and  established  the  fact  that  the  fat  of  the  cells  is 
split  into  its  components,  and  that  the  fatty  acids  combine  (at  least  in 
part)  with  calcium.  Dettmer  "  found  that,  although  fresh  pancreatic 
juice  caused  fat  necrosis,  a  commercial  preparation  of  trypsin  did 
not  do  so,  and,  therefore,  he  concluded  that  probably  the  lipase  of  the 

82  See  Miiller,  Cent.  inn.  Med.,  1907   (28),  2!)7. 

82a  Jour.  Exp.  I\Icd.,  1914    (19),  239;   Zoit.  Immunitat.,  1914    (23),  71. 

83  General  literature  will  be  found  in  tlie  articles  eited  in  tlie  text :  also  in 
Opie's  "Diseases  of  the  Pancreas";  and  in  Truhart's  "Pankreas-Patliolojiie." 
Wiesbaden,   1902. 

84  The  fatty  acids  form  masses  of  crystals  in  the  fat-cells,  and  they  can  also 
be  demonstrated  microcheniically  by  Penda's  method  (Virchow's  Arch.,  1900  (  Ifil), 
194),  which  ('(msists  of  staininfjr  with  a  copper  acetate  mixture,  blue-firccii  copper 
salts  of  the  fattv  acids  beini;  formed. 

K-'WuHV  (l?eri.  klin.  Woch.,  1902  (39),  734)  claims  to  have  observed  an  excep- 
tion.to  this  rule,  l)ut  liis  accoimt  is  not  by  itself  convincinii.  Fabyan  (.lohns 
Hoi)kins  llosp.  H>ill.,  1907  (IS),  349)  reports  a  case  of  multiple  subcutaneous 
fat  necrosis  witliout  pancreatic  lesions,  in  a  14  days'  old  l)aby,  and  j^ivcs  a  re- 
view of  other  similar  cases. 

8«  Virchow's  Arch.,  1H90   (122),  2r)2. 

8T  Dissertation,  Gottingen,  1895. 


FAT  NECROSIS  385 

pancreatic  juice  was  the  active  agent.  Flexner  ^^  supported  this  con- 
tention by  demonstratinfj:  tlie  presence  of  a  fat-splitting  enzyme  in 
foci  of  fat  necrosis,  wliich  was  corroborated  by  Opie.-^  The  latter'*" 
was  also  able  to  (lenioiistrate  the  presence  of  lipase  in  the  urine  of  a 
patient  with  fat  necrosis,'"  and  the  highest  values  for  amylase  in  the 
blood  and  urine  are  found  in  pancreatitis  (Stocks).'*^'' 

In  a  study  of  the  pathogenesis  of  fat  necrosis,  particularly  with 
reference  to  the  question  whether  the  lipase  or  the  trypsin  of  the 
pancreatic  juice  was  responsible.  Wells  ^-  found  that  typical  fat 
necrosis  could  be  produced  by  injecting  extracts  of  fresh  pancreas 
into  animals,  either  of  the  same  species  as  that  from  which  the  pan- 
creas was  obtained,  or  into  a  foreign  species.  Commercial  "pan- 
ereatins"  were  also  quite  effective,  whether  in  weak  acetic  acid  or 
weak  alkaline  solutions.  The  power  of  these  materials  to  cause  fat 
necrosis  was  reduced  by  heating  to  or  above  60°  for  five  minutes, 
and  completely  destroyed  at  71°,  indicating  that  the  active  agent 
is  an  enzyme.  But,  as  in  the  same  material  trypsin  was  injured  by 
temperatures  above  60°,  and  destro.yed  at  between  70°  and  72°,  and 
lipase  was  weakened  above  50°,  and  destroyed  above  70°,  it  was  im- 
possible to  determine,  by  heating  pancreatic  preparations,  whether 
the  lipase  or  the  trypsin  was  the  essential  factor.  By  permitting 
pancreatic  extracts  to  digest  themselves  it  was  found  that  the  power 
to  produce  fat  necrosis  decreased,  pari  passu,  with  the  decrease  in 
lipolytic  strength.  Preparations  strongly  tryptic,  but  very  weak  in 
lipase,  produced  no  fat  necrosis,  and,  on  the  other  hand,  extracts  of 
pig's  liver  or  of  cat's  serum,  both  rich  in  lipase  but  devoid  of  tryp- 
sin, were  equally  ineflPective.  Furthermore,  mixtures  of  liver  or 
serum  lipase  and  trypsin  were  incapable  of  causing  fat  necrosis. 
Fresh  pancreatic  extracts  from  fasting  dogs,  containing  lipase  but 
almost  no  trypsin  (which  in  fresh  extracts  is  still  in  the  form  of 
inactive  trypsinogen),  produced  abundant  fat  necrosis,  whereas  after 
the  trypsinogen  in  such  extracts  was  activated  by  enterokinase,  no 
fat  necrosis  could  be  produced.  It  therefore  seems  certain  that 
trypsin  alone  cannot  produce  fat  necrosis,  and  that  the  decrease  in 
strength  of  lipase  in  a  pancreatic  extract  is  associated  with  a  cor- 
responding decrease  in  power  to  produce  fat  necrosis.  But,  on  the 
other  hand,  lipase  of  liver  or  blood-serum  alone,  or  when  mixed  with 

88  Jour.  Exper.  Med.,  1807   (2),  413. 

89Contrib.  of  pupils  of  W.  IT.  Welch,  Baltimore.  1000,  p.  850;  .Tolnis  Hopkins 
Hosp.   Rep.,    1000    (0),   8.50. 

00  Opie,  "Diseases  of  the  Pancreas,"  Lippincott,  1003,  p.  ].")6:  .Johns  Ildiikins 
Hosp.  Bull.,   1002    (13),  117. 

91  It  yet  remains  to  be  seen  if  this  is  a  constant  occurrence:  and  also  if  tlie 
lipase  so  excreted  comes  from  the  pancreas,  for  Zeri  (II  Policlinico.  100.5  (12), 
"3.3)  has  found  lipase  in  the  urine  in  hcmorrhajric  nepliritis  and  inihimiiiation  of 
the  urinary  tract:  also  Pribram  and  Loewv,  Zeit.  phvsiol.  Chem.,  1012    (76),  480 

9ia  Quart.  .lour.  Med.,  1016   (0),  216. 

92  Jour.  Med.  Research,  1003   (9),  70. 

25 


386  RETROGRESSIVE    CHANGES 

trypsin,  will  not  produce  fat  necrosis.  The  possibility  remains  that 
pancreatic  lipase  is  different  from  liver  or  serum  lipase,  and  can 
by  itself  cause  fat  necrosis;  more  probably,  however,  the  production 
of  fat  necrosis  depends  upon  a  double  action,  tr\'psin  causing  the 
death  of  the  cells,  and  lipase  splitting  the  fats."^  The  fatty 
acids  alone  will  not  cause  necrosis  of  fat-cells,  and  it  was  shown 
that  the  first  steps  in  the  process  consist  of  a  necrosis  of  the  surface 
endothelium  extending  into  the  connective  and  fat  tissue;  this  may 
occur  in  a  few  minutes,  while  evidence  of  fat-splitting  can  be  ob- 
tained onlj^  after  about  three  hours,  and  the  splitting  occurs  only  in 
cells  that  have  already  become  necrotic ;  hence  the  fat-splitting  is 
not  the  cause  of  the  necrosis,  but  occurs  subsequent  to  the  necrosis. 
After  about  four  hours  a  substance  appears  in  the  decomposed  fat 
that  stains  with  hematoxylin,  which  is  probably  calcium. 

Fat  necrosis  may  be  produced  by  any  means  that  will  cause  the 
escape  of  pancreatic  juice  from  the  natural  channels  within  the 
gland.  In  human  pathology  it  has  followed  trauma  and  acute  in- 
fection of  the  gland,  and  the  blocking  of  the  ampulla  of  Vater  b^^  gall- 
stones which  permits  the  bile  to  back  up  into  the  pancreatic  duct, 
where  it  produces  an  acute  inflammation  of  the  pancreas  (Opie)."* 
Flexner  °^  has  shown  that  it  is  the  bile  salts  that  cause  the  inflamma- 
tion, and  also  that  this  effect  is  decreased  or  prevented  by  the  presence 
of  large  amounts  of  colloids.  ]\Iuch  emphasis  is  laid  by  some  au- 
thors ^**  upon  the  necessity  of  enterokinase  passing  up  the  ducts  to 
activate  the  trypsinogen  (an  idea  first  advanced  by  Starling  and  Ba}^- 
Hss  in  1902),  but  it  should  be  remembered  that  there  are  kinases  pres- 
ent in  leucocytes,  and  that  kinases  can  develop  in  the  pancreas  itself 
during  autolysis,  which  can  activate  the  trypsinogen ;  hence  the  pres- 
ence of  entero-kmase  is  not  essential  for  sufficient  activation  of  tryp- 
sinogen to  account  for  pancreatitis  and  fat  necrosis.  Lattes  °'^  believes 
that  fresh  pancreatic  juice,  which  digests  tissues  very'  slowly,  can  pro- 
duce typical  fat  necrosis  but  not  the  characteristic  intoxication;  this 
results  from  the  action  of  juice  which  has  been  activated  by  entero- 
kinase, or  by  products  of  pancreatic  autolysis  M'hicli  have  a  similar 
effect.     The  kinases  of  leucocytes  he  found  unable  to  activate  pan- 

!i3  Wlion  fat  tissue  dies  in  the  liody  from  other  causes,  tlie  lipase  normally  eon- 
tain<'d  within  the  fat  tissue  does  not  cause  the  changes  seen  in  fat  necrosis.  It  is 
possible,  therefore,  that  the  coml)inin<r  of  newly  split  fatty  acids  hy  the  alkali  of 
tlie  j)ancreatic  juice  is  resjjonsihle  for  tlie  formation  of  the  larpe  amount  of 
soajjs  found  in  fat  necrosis.  Otheiw  isc  we  mipht  exjx'ct  the  lijjase  to  ])roduee  only 
an  ('(piilihiium,  and  tliat,  in  tlie  case  of  fat,  s(^eins  to  exist  wlien  most  of  the 
substance  is  neutral  fat.  In  support  of  this  idea  I  foiuid  that  stronix  alkalies 
injected  into  fat  tissue  sometimes  caused  changes  very  closely  resembling  areas 
of  fat  necrosis  in  the  early  st,ages. 

»4Bull.  dohns  Hopkins  'llosp.,  1001    (12),  182. 

o'-.  Jour.  Exp.  Med.,  lOOG    (8),  107. 

0"  I'olva,  ]\1itt.  Grenz.  Med.  u.  Chir.,  1911  (24),  1;  Rosenbach.  Arch.  klin.  (hir., 
1910    (93),  278. 

07  Virchow'a  Arch.,  1913   (211),  1. 


FAT  NECIiOSIS  387 

ereatic  trypsinojren  sufficiently  to  make  it  highly  toxic  These  ob- 
servations indicate  that  in  pancreatic  necrosis  it  is  the  kinase  liberated 
from  the  autoly/.in<r  necrotic  tissue  -wliich  is  responsil)]e  for  the  acti- 
vation and  resultin":  toxic  effects  of  the  trypsinopren.  As  a  result  of 
injury  by  bile  salts,  or  any  other  agent  that  produces  cell  death,  the 
(lead  and  injured  cells  are  digested  by  the  pancreatic  juice  which  is 
thus  further  activated  and  makes  its  escape  into  the  surrounding  fat 
tissue.  Wells'  experiments  showed  that  the  lesions  of  fat  necrosis 
may  be  produced  in  three  to  five  hours,  large  enough  to  be  visible  to 
the  naked  eye ;  their  form  and  size  depend  solely  upon  the  area  of  fat 
tissue  exposed  to  the  action  of  the  pancreatic  juice.  The  process 
progresses  for  but  a  few  hours,  the  extension  seeming  to  be  limited  by 
surrounding  leucocytes.  The  lesions  may  appear  at  remote  points  in 
the  thoracic  and  pericardial  cavities  or  in  the  subcutaneous  tissues, 
the  causative  agent  probably  being  canned  by  the  lymphatic  vessels, 
possibly  in  the  form  of  emboli  of  pancreas  cells.^  There  may  even  be 
some  splitting  of  the  fats  in  the  liver  in  these  cases,  with  intrahepatic 
necrosis.-  Fat  necrosis  itself  is  not  dangerous  to  the  affected  organ- 
ism, the  associated  pancreatitis  fand  peritonitis)  causing  all  the 
symptouis.^  There  is  no  evidence  that  sufficient  quantities  of  soaps 
(which  are  toxic)  are  absorbed  from  the  necrotic  areas  to  cause  appre- 
ciable intoxication.  The  soaps  that  are  formed  in  the  necrotic  areas, 
indeed,  are  probably  not  much  absorbed,  but  are  precipitated  as  cal- 
cium soaps ;  in  such  areas  at  least  as  high  as  85  per  cent,  of  the  soaps 
may  be  insoluble.*  Healing  follows  rapidly  in  case  of  recovery ;  the 
foci  may  disappear  as  early  as  eleven  days  after  their  formation  (in 
experimental  animals). 

In  the  urine  of  persons  with  pancreatitis  is  frequently  found  a 
substance  forming  an  osazone,  which  has  been  the  subject  of  much 
investigation   because   of   its  possible   diagnostic   value.     Cammidge,^ 

iPavr  and  Martina.  Dent.  Zeit.  Chir.,  1006   (F.3),  189. 

2  See  Berner.  Virchow's  Arch.,  1007    (187),  .300. 

3  Giileke  (Arch.  klin.  Chir.,  1008  (8,5),  015)  considers  the  intoxication  of  acute 
pancreatitis  as  an  intoxication  Avith  trvpsin.  A\liich  can  be  checlced  bv  antitrAjisin. 
Doberauer  (Beitr.  klin.  Chir..  lOOG  (48),  450),  Esdahl  (.Tour.  Exp.  Med.'.  1007 
(0).  .385),  and  Petersen,  .Tol)lin£r,  and  Egprstein.  ibid.,  1016  (23),  401,  however, 
look  upon  tlie  products  of  cellular  disintegration  as  the  source  of  tlie  intoxica- 
tion. V.  Bercruiann  (Zeit.  exp.  Path.  u.  Ther..  1006  (3),  401,  states  that  the 
toxicity  is  not  due  to  either  tlie  enzymes  or  to  allnimoses;  and  that  it  is  a  true 
autointoxication  Avhich  can  be  prevented  by  ^irevious  immunization  witli  either 
pancreas  extracts  or  commercial  trvpsin.  (See  also  Fischler,  Deut.  Arch.  klin. 
Med.,  1011  (103).  156:  and  v.  Bergmann  and  Culeke,  IMiinch.  med.  Woch.,  1010 
(57),  1673).  The  histones  and  protamines  liberated  from  the  digested  tissue, 
and  which  are  very  toxic,  have  been  sugprested  as  a  possible  factor  by  Schitten- 
helm  and  Weichardt  (Zeit.  Immunitiit..  1013  (14),  600),  while  the  beta-nucleo- 
proteins  are  inchided  among  the  toxic  elements  bv  Goodpasture.  Jour.  Exp.  Med.. 
1017   (25),  277. 

4  See  Frugoni  and  Stradiotti,  Arch.  Sci.  !Med.,  Torino,  lOlO;  also  Berl.  klin. 
Woch..  1010   (47).  386. 

sProc.  Royal  Soc.  Med.,  1010  (III,  pt.  2),  103,  bibliographv:  Lancet,  1014, 
Sept.  26.  ... 


388  RETROGRESSIVE    CHANGES 

who  first  described  a  reaction  based  on  this  observation,  considers 
that  the  substance  is  a  pentose,*^  derived  from  the  nucleoproteins  of  the 
pancreas;  it  bears  no  relation  to  the  fat  necrosis,  but  is  commonly 
found  with  fat  necrosis  because  of  the  associated  pancreatitis.  Pre- 
sumably cell  necrosis  elsewhere  than  in  the  pancreas  may  at  times 
cause  the  same  reaction  to  appear/  In  pancreatitis  with  fat  necrosis, 
or  whenever  there  is  any  injury  to  the  pancreas,  there  may  be  found 
an  increase  in  the  amount  of  diastase  in  the  blood  and  urine,  sufficient 
to  be  of  diagnostic  value  according  to  Y.  Noguchi.'^^  The  peritoneal 
exudate  in  acute  pancreatitis  is  not  toxic,  contains  no  free  trypsin  and 
is  no  more  lipolytic  than  normal  serum,  presumably  because  of  neu- 
tralization of  the  enzymes  and  poisons  by  the  exuded  plasma.'" 

Self -digestion  of  the  pancreas  occurs  soon  after  death,  and  the  pan- 
creatic juice  may  in  this  way  bring  about  a  portmortem  fat  digestion 
that  resembles  somewhat  the  intravital  fat  necrosis  in  its  gross  ap- 
pearances,® and  Wells  found  that  the  same  changes  might  be  pro- 
duced by  injecting  pancreatin  into  the  bodies  of  dead  animals,  or 
by  keeping  fat  tissue  in  pancreatin  solutions.  Wulff  found  that  fatty 
acids  were  demonstrable  by  Benda's  method  in  the  pancreas  of  nearly 
all  cadavers.  The  process  differs  from  the  intra  vitam  form  in  being 
less  sharply  circumscribed,  and  microscopically  by  the  absence  of  cellu- 
lar and  vascular  reaction.  That  the  essential  changes  of  fat  necrosis 
can  be  produced  postmortem  is  final  proof  that  they  are  due  to 
enzymes,  rather  than  to  circulatory  or  cellular  action. 

GANGRENE 

This  term  indicates  merely  that  certain  marked  secondary  changes, 
either  putrefaction  or  desiccation,  have  occurred  in  necrotic  areas  of 
some  size.  Hence  we  have  the  chemical  changes  of  putrefaction 
added  to  those  of  necrosis  in  the  case  of  moist  gangrene,  whereas  in 
dry  gangrene  nearly  all  the  chemical  changes  are  brought  to  a  stand- 
still through  the  desiccation.  In  the  latter  it  is  only  at  the  line  of 
demarcation,  where  some  moisture  remains,  that  chemical  changes 
still  go  on ;  these  consist  chiefly  of  autolysis  of  the  dead  tissues,  and 
also  of  their  digestion  by  leucocytes,  which  results  eventually  in  the 
separation  of  the  dead  tissue  from  the  living;  this  is  best  seen  after 
surface  burns,  carbolic-acid  gangrene,  etc. 

Moist  gangrene  is  accompanied  by  the  dual  action  of  the  cellular 
enzymes  and  of  the  putrefactive  organisms  that  are  growing  in  the 

c  Weber  believes  that  it  is  a  liexose  (Dent.  med.  Wocli.,  ini2  (3S),  106).  and 
it  may  be  urinary  dextrin  (Pekelliaring  and  Van  IToo>renhuyze,  Zeit.  phvsiol. 
Chem."   1014    (91),'  151). 

7  See  Wliipple,  et  al,  Johns  Hopkins  TIosp.  ■Bull..  1010  (-21).  XV.) -.  Karas.  Zeit. 
klin.  Med.,  1013   (77).  125. 

7a  Arch.  klin.  C'hir.,  1012   (OS).  TT.  2. 

7b  Whipple  and  Coodpastiire,  Siir";..  Gvn.  and  OI)st..  lOi;?    (17),  541. 

sChiari,  Zeit.  f.  Ileilk.,  180(1  (17),  00 ;"  Pforrinjrer,  Virehow's  Arch..  ISOO  (15S), 
126;  Liepmann,  ibid.,  1002   (100),  532;  WulfT,  Bcrl.  klin.  Woch.,  1002    (30),  734. 


GANGRENE  389 

(lead  tissue,  and  as  a  result  such  tissue  contains  all  the  innumerable 
products  of  the  decomposition  of  proteins  and  fats.  Thus  Ziegler 
mentions  as  morphological  elements  that  may  be  present  in  gan- 
grenous tissue:  Fat  needles,  the  so-called  "margarin"  crystals  (a 
mixture  of  stearic  and  palmitic  acids),  fine  acicular  crystals  of 
tyrosine,  globules  of  leucine,  rhombic  plates  of  triple  phosphate,  black 
and  brown  nuisses  of  pigment,  and  crystals  of  hematoidin.  In  solu- 
tion we  also  have,  beyond  a  doubt,  all  the  substances  formed  in  the 
decomposition  of  proteins,  from  proteoses  and  peptones  down  through 
the  different  amino-acids  to  such  final  products  as  ammonia  and  its 
salts,  while  CO.^  and  1L,S  are  abundantly  given  off.  In  addition 
occur,  undoubtedly,  many  of  the  ptomains  which  are  formed  by  the 
action  of  the  bacteria  upon  the  amino-acids  derived  from  the  pro- 
teins." In  the  sputum  from  pulmonary  gangrene  there  is  but  little 
soluble  protein,  most  of  the  nitrogen,  of  which  there  is  much,  is  in 
the  formed  elements. ^'^  The  fetid  plugs  which  occur  in  the  bron- 
chioles in  gangrene,  the  "Dittrich's  plugs,"  were  found  by  Traube 
to  be  composed  chiefly  of  fatty  acid  crystals,  and  Schwartz  and 
Kayser  ^^  ascribe  their  formation  to  the  action  of  lipolytic  staphy- 
lococci. 

If  the  necrotic  tissue  is  in  contact  with  living  tissue  over  a  con- 
siderable area,  enough  of  these  products  of  autolysis  and  putrefaction 
may  be  absorbed  to  cause  intoxication  (sapremia) .  At  the  same  time, 
the  formation  of  such  large  quantities  of  crystalloids  from  the  pro- 
teins of  the  dead  tissue  leads  to  a  dififusion  of  water  into  this  area, 
with  consequent  swelling,  and  often  a  lifting  up  of  the  skin  in  the 
form  of  blisters. 

Emphysematous  gangrene,^-  usually  produced  by  gas-forming 
anaerobic  bacteria,  particularly  by  B.  aerogenes  capsulatus,  may  also 
possibly  be  produced  by  B.  coli  communis  in  diabetic  patients  in 
whose  blood  and  tissues  there  may  occur  sufficient  sugar  to  permit  of 
gas-formation.  Ilitsehmann  and  Lindenthal  ^^  found  that  the  gas 
produced  in  cultures  by  an  anaerobic  organism  which  they  isolated 
from  a  case  of  emphysematous  gangrene,  consisted  of  67.55  per  cent, 
hydrogen,  30.62  per  cent,  carbon  dioxide,  and  traces  of  ammonia 
and  nitrogen ;  this  corresponds  to  the  statement  of  Welch  and  Nut- 
tall  that  the  gas  in  the  tissues  of  infected  animals  is  inflammable. 
Dunham  ^*  found  that  the  gas  produced  by  B.  aerogenes  capsulatus  in 
cultures  has  the  following  composition :     Hydrogen,  64.3  per  cent. ; 

9  An  interestinor  observation  concerning:  gangrene  of  the  lung  has  been  made 
by  Eijkman  (Cent.  f.  Bakt.,  Abt.  1,  1003  (35),  1),  who  found  in  this  condition 
bacteria  that  secrete  an  enzyme  dissolving  elastic  tissue. 

loOrszag.  Zeit.  klin.  Med.,'  1909   (67),  204. 

11  Zeit.  klin.  Med.,  1905    (5G),  111. 

12  Complete  literature  bv  Fraenkel,  Ergebnisse  der  Pathol.,  1902  (8),  403;  and 
by  Welch,  Johns  Hopkins  Hosp.  Bull.,  1900   (11),  185. 

13  Quoted  by  Fraenkel. 

14  Johns  Hopkins  Hosp.  Bull.,  1897   (8),  68. 


390  RETROGRESSIVE    CHANGES 

carbon  dioxide,  27.6  per  cent. ;  other  gases,  probably  chiefly  nitrogen, 
8.1  per  cent. 

RIGOR  MORTIS  15 

This  topic  may  be  appropriately  considered  in  connection  with  cell 
death,  since  it  is  a  characteristic  change  occurring  after  general 
death.  All  forms  of  muscle,  striped,  smooth,  and  cardiac,  undergo 
this  change,  which  is  shown  by  a  shortening  and  thickening  of  the 
muscle,  which  also  becomes  opaque  and  hard.  Rigor  mortis  begins 
first  in  the  heart  muscle,  according  to  Fuchs,^''  but  it  is  generally 
observed  first  in  the  eyelids,  then  in  the  muscles  of  the  jaw,  from 
which  point  it  proceeds  downward,  although  the  ujjper  extremities 
may  not  become  rigid  before  the  lower.  The  time  of  onset  is  ex- 
iremely  variable,  but  the  following  general  rules  may  be  stated :  All 
conditions  that  lead  to  excessive  muscular  metabolism,  with  its  re- 
sulting increase  in  the  acidity  of  the  muscle  fluids,  will  hasten  the 
onset  of  rigor  mortis ;  thus,  people  killed  suddenly  during  violent 
activity  may  remain  almost  in  the  position  in  which  they  met  death. 
Acute  fevers,  strychnine  poisoning,  tetanus,  etc.,  cause  likewise  a 
rapid  onset  of  rigor,  which  may,  indeed,  appear  almost  simultane- 
ously with  death  or  even  before  the  heart  has  stopped  beating. 
When  a  liealthy  individual  meets  death  without  previous  exertion, 
rigor  does  not  usually  appear  for  four  or  six  hours,  but  will  be 
hastened  by  heat  and  retarded  by  cold.  Death  from  hemorrhage  or 
asphyxia  is  followed  by  a  slow  development  of  the  rigor.  Under 
ordinary  conditions  rigor  usually  begins  between  the  first  and  second 
hour  after  death  and  is  complete  in  one  or  two  more  hours. ^' 

The  duration  of  rigor  mortis  also  is  influenced  by  many  factors. 
In  general,  it  may  be  said  that  the  duration  is  in  inverse  relation 
to  the  rapidity  of  onset,  and  directly  to  the  musculature  of  the  in- 
dividual. Therefore,  in  an  emaciated  individual  dying  with  fever, 
rigor  may  appear  and  disappear  again  within  two  or  three  hours,  or, 
indeed,  escape  observation  altogether.  The  body  of  a  muscular  man 
dying  from  accident  or  hemorrhage  may,  on  the  other  hand,  show 
rigor  for  two  or  three  weeks  if  kept  in  a  cold  place.  Once  the  rigor 
has  been  broken  by  force,  it  does  not  again  return. 

Rigor  mortis  may  be  produced  even  before  death,  through  poisons 
(monobromacetic  acid,  quinine),  and  its  occurrence,  even  postmor- 
tem, does  not  necessarily  mean  that  the  nniscle  is  dead,  for  if  the  part 
is  transfused  with  a  salt  solution  the  rigor  may  be  removed,  and  the 

15  Literature,  see  v.  Fiirlli,  TTini(ll)ucli  d.  Tlioclipm.,  100!)  (II  (2),  252;  also 
Molt/cr  and  Aiicr,  Jour.  Kxp.  Med.,   IltOS    (10),  45). 

inZeit.  f.  Ileilk.,  IHOO   (21,  Patli.  Abt.),  1. 

17  Kifror  mortis  may  develop  in  tlie  dead  fetus  wliile  in  the  womb,  but  it  cen- 
erallv  disajipears  witliin  live  or  six  lioiirs.  Literature  hv  WolfT,  Areli.  f.  Gvn., 
]!)0:}'  (G8),  549;  Das,  Brit.  Jour,  of  Obstet.,  1903    (4),  545. 


RIGOR  MORTIS  391 

muscle  will  thou  be  found  to  react  to  stimuli.  This  indicates  that  the 
chemical  chano:es  of  rigor  mortis  are  not  very  profound.^** 

The  chemistry  of  the  changes  involved  in  rigor  mortis  has  been 
a  much-contested  problem.  Two  chief  doctrines  have  been  sup- 
ported :  one  that  rigor  was  not  essentially  different  from  ordinary 
muscular  contraction  except  in  degree,  and  perhaps  due  to  a  loss 
of  inhibition  to  contraction.  The  other  looks  upon  it  as  a  coagula- 
tion similar  to  the  coagulation  of  the  blood ;  and  this  idea,  it  may  be 
said,  has  had  the  most  general  acceptance.  Brlicke  in  1842  supported 
this  view,  and  in  1859  Kiihne  extracted  from  muscle  a  plasjua  which 
coagulated  like  ordinary  blood  plasma.  The  protein  which  formed 
the  clot  is  called  myosin,  and  its  coagulable  antecedent,  myosinogen. 

This  experiment  ha.s  been  since  repeatedly  verified  and  amplified, 
especiall}'  by  v.  Fiirth  and  by  Halliburton,"  who  have  separated 
more  definitely  the  proteins  concerned  in  coagulation,  and  found 
them  to  be  globulins.  There  seem  to  be  two :  one,  coagulating  at  47°, 
called  paramyosinogen  (Halliburton),  constitutes  but  about  one-fifth 
of  the  total  clotting  globulin,  and  passes  readily  into  the  insoluble 
clot,  myosin:  the  other,  which  coagulates  at  56°,  constitutes  the  re- 
maining four-fifths,  is  called  myosinogen  (Halliburton),  or  myogen 
(v.  Fiirth),  and  before  becoming  changed  into  myosin  it  passes 
through  a  soluble  stage  called  soluble  myogen-fibrin,  which  is  coagu- 
lated at  the  remarkably  low  temperature  of  40°. 

By  analogy  with  fibrin-formation  we  should  expect  this  clotting 
also  to  be  brought  about  by  an  enzyme,  but  this  has  not  been  proved. 
Calcium  is  of  influence,  favoring  coagulation  greatly,  but  its  presence 
is  not  absolutely  essential  (v.  Fiirth).  Of  particular  importance  is 
the  acid  reaction  of  the  dead  muscle.  Normal  muscle  is  amphoteric 
when  at  rest,  but  when  active  the  reaction  becomes  more  and  more 
acid,  as  it  also  does  when  the  circulation  is  shut  off,  and  hence  acidity 
increases  greatly  after  death.  The  acidity  is  due  chiefly  to  lactic 
acid  (although  the  neutral  phosphates  may  become  converted  into 
acid  phosphates  in  the  presence  of  the  lactic  acid,  and  thus  seem 
to  contribute  to  the  acidity),  and  may  increase  in  twenty-four  hours 
after  death  by  from  6.7  to  12.8  c.c.  of  "/^o  acid  for  each  100  grams 
of  muscle  (v.  Fiirth -°).  The  same  author  found  that  although  the 
amount  of  acid  might  become  in  time  sufScient  to  cause  coagula- 
tion of  the  muscle  proteins  by  itself,  yet  actually  rigor  mortis  appears 
before  the  acidity  has  reached  any  such  degree.  Meigs  -^  advanced 
the  hypothesis  that  the  rigor  is  due  to  the  swelling  of  the  muscle  col- 
loids under  the  influence  of  acids,  a  view  which  is  accepted  by  von 

isSoe  Mangold.  Pfliiper':,  Arch.,   1003    (Ofi),  4nS. 

19  "Chemistry  of  ^fuscle  and  Xerve."  1004. 

20  Hofnieister's  Beitr.,  1003  (3),  r)43 :  see  also  Fletcher  and  Hopkins,  Jour,  of 
Phvsiol..   1007    (3.5),  247;   Wacker.  Biochem.  Zeit.,   1016    (75),   101. 

2iAmer.  Jour.  Physiol.,  1010   (26),  101. 


392  RETROGRESSIVE    CHANGES 

Fiirtli  and  Lenk.--  When  sufficient  acid  is  formed  in  the  muscle 
the  swelling  may  be  so  great  that  the  structure  of  the  muscle  cell  is 
destroyed  entirely,  and  it  goes  into  the  condition  of  "waxj-  degenera- 
tion."-'^ This  readih'  explains  why  the  time  of  appearance  of  rigor 
is  so  modified  by  the  amount  of  muscle  metabolism  before  death.  It 
is,  indeed,  possible  to  produce  rigor  in  living  animals  by  transfusing 
a  limb  with  slightly  acid  salt  solution,-*  and  in  strychnine-poisoning 
the  muscular  spasm  may  pass  imperceptibly  into  rigor  mortis. 

It  has  been  suggested  that  the  disappearance  of  rigor  mortis  depends 
upon  beginning  autolj'sis  of  the  clot  by  the  intracellular  proteases  of 
the  muscle,  which  act  best  in  an  acid  medium,  but  proteoses  and  pep- 
tones cannot  be  found  in  such  muscle.  It  is  improbable  that  the  de- 
gree of  acidity  ever  becomes  so  high  that  the  myosin  is  redissolved 
through  a  conversion  into  acid  albumin  (syntonin),  as  was  formerly 
supposed.  V.  Fiirth  holds  that  the  re-solution  of  the  rigor  is  caused 
by  coagulation  of  the  proteins,  thus  reducing  this  hydrophilic  tend- 
ency, a  view  in  harmony  with  recent  developments  in  colloid  chemis- 
try.-^ 

"Waxy"  degeneration  of  muscles,  although  usually  resulting  from 
the  action  of  toxic  substances,  is  entirely  different  from  cloudj-  swell- 
ing, in  that  the  cytoplasm  has  become  homogeneous  and  not  granular. 
This  is  undoubtedly  due  to  the  increased  accumulation  of  acid  which 
takes  place  in  muscles  when  they  suffer  from  a  defective  oxygen  sup- 
ply, for  I  have  found  it  possible  to  produce  the  tj'pical  appearance 
of  Zenker's  waxy  degeneration  by  letting  weak  solutions  of  lactic 
or  other  acids  act  on  muscle  fibers.-^^  Even  excessive  stimulation  of 
muscles  was  found  to  be  sufficient  to  cause  waxy  degeneration,  the 
acid  being  formed  faster  than  it  can  be  removed.-*^ 

INIuscles  showing  the  ' '  reaction  of  degeneration ' '  have  been  analyzed 
by  Rumpf  and  Schumm,-^  who  found  a  great  increase  in  the  fatty 
matter,  which  was  about  fifteen  times  the  normal  amount.  The  muscle^ 
deducting  the  fat,  showed  a  loss  of  solid  matter  and  an  increase  of 
water;  sodium  and  calcium  were  increased,  potassium  decreased. 
There  is  also  a  great  relative  increase  in  the  proportion  of  phosphorus. 

22Biochcm.  Zeit.,  1011    (33),  341;  Wien.  klin.  Woch.,  1!U1    (24),  1070. 

23  Wells,  Jour.  Exper.  Med.,  1000   (11),  1. 

24  The  hardness  of  a  linili  from  which  the  blood-supply  has  been  shut  ofl'  by 
tlirombosis  or  einbolisin,  and  also  much  of  the  cramp-like  i)ain.  is  probably  due 
to  rijror  mortis  in  the  muscles  caused  by  acid  formation  under  conditions  of 
sub-oxidation. 

25  Corroborated  bv  Lentz,  Zeit.  anpew.  Chem.,  1012  (25),  1513:  and  Schwarz, 
Biochem.  Zeit.,   1012    (37),  35. 

2'ia  Jour.  Exper.  ]\Icd.,  1000  (11),  1.  Corroborated  bv  Steinmler,  \irc]io\v"s 
Arch.,   1014    (21fi),  57. 

2"  As  this  work  antedates  much  of  the  recent  work  on  the  influence  of  acids  of 
metabolic  origin  upon  the  swelling  of  cell  structures,  attention  nuiy  be  called  to 
the  fact  that  a  preliminary  report  of  these  experiments  was  made  in  the  first, 
edition  of  this  book,  written  in  100(i. 

27  Deut.  Zeit.  f.  Nervenheilk.,  1001    (20),  445. 


ATROPHY  393 

bound  to  })r()teiii  in  imisrlcs  wiiicli  have  atrophied  after  nerve  section, 
because  of  the  jx'i-sistenee  of  iiuelear  and  loss  of  non-nueh'ar  elements 
(.Grund -"''),  but  there  is  little  ehanjie  in  the  i^roportion  of  mono-  and 
di-amino  nitrogen.^^ 

ATROPHY 

The  chemical  chanpfes  of  simple  atroi)liy  have  not,  so  far  as  I  can 
find,  been  definitely  studied.  It  is  to  be  presumed,  in  view  of  the 
structural  changes,  that  analysis  of  atrophied  tissues  would  show  a 
relatively  high  nucleic  acid  and  collagen  content.  It  is  known  that  in 
atrophy  the  cell  lipoids  are  not  much  altered,  while  the  simi)ler  fats 
may  be  increased  in  parenchymatous  organs.  In  fatty  tissues,  of 
course,  the  fat  is  greatly  reduced,  its  place  being  partly  taken  by 
serum  (serous  atrophy  of  fat).  In  the  heart  muscle,  especially,  but 
also  to  a  less  extent  in  the  liver  and  kidney,  during  atrophy  there  is  an 
increased  pigmentation  (brown  atrophy)  apparently  consisting  of 
lipochromes  or  lipofuscins ;  but  it  is  to  be  doubted  that  this  represents 
so  much  an  actual  increase  in  pigment  as  a  relative  increase  through 
loss  of  other  cellular  elements.  Atrophied  tissues  also  tend  to  undergo 
a  marked  compensatory  invasion  by  fatty  areolar  tissue  if  located  in 
contact  with  such  tissue ;  e.  g.,  atrophy  of  muscles  after  nerve  section, 
specific  muscular  dystrophies,  and  atrophy  of  the  pancreas. 

Starvation,  of  course,  produces  typical  atrophic  changes  in  the 
tissues,  and  the  general  effects  on  metabolism  have  been  especially 
fully  worked  out  by  Benedict.-'*-''  The  structural  changes  in  parenchy- 
matous cells  are  described  ~^^  as  of  two  types ;  first,  granular  changes 
and  vacuolization  of  the  cytoplasm,  resembling  the  effects  of  osmotic 
pressure  alterations;  second  and  later,  lysis  of  cytoplasm  with  also 
some  involvement  of  the  nuclei,  after  the  order  of  autolytic  changes. 
The  cell  walls  may  also  become  indistinct,  so  that  the  cells  resemble  a 
syncytium. -*'°  In  the  atrophied  muscle  after  nerve  section  Wake- 
man  ^^^  found  a  decrease  in  solids,  and  a  lowered  proportion  of 
diamino  acids. 

Morse  has  considered,  by  experimental  methods,  the  question  of  the 
mechanism  involved  in  atrophy,  using  especially  the  involuting  tail 
of  the  tadpole  as  his  test  object.-^®  He  could  find  no  evidence  that 
autolysis  is  accelerated  during  this  involution,  nor  in  the  atrophying 
muscle  after  nerve  section.  The  involution  of  the  puerperal  uterus, 
whether  it  can  properly  be  called  atrophy  or  not,  seems  to  be  the 
result  of  heightened  autolysis,  the  products  of  which  are  excreted 

2TaArch.  exp.  Path.,  1912    (67),  .39.3. 

28Wakeman,  .Tour.  Biol.  Chem.,  190S   (4),  r37. 

28aCarnefrie  Inst.  Piihl.,  1915,  No.  203. 

28b  Cesa-Biandii.  Frankf.  Zeit.  Path.,  1909    (3).  723. 

28cMorgulis,  Howe  and  Hawk,  Biol.  Bull.,  1915   (28),  397, 

28dJour.  Biol.  Chem.,  1908    (4),  137. 

28eAmer.  Jour.  Physiol.,  1915    (36),  145. 


394  ri:tro(:ri:s><i\  i:  cjLwaES 

quantitatively  in  the  urine. -^^  Bradley  -^^  calls  attention  to  the  fact 
that  atrophy  occurs  commonly  under  conditions  of  reduced  blood 
supply,  Avhich  implies  i^artial  asphyxia  and  a  resulting  tendency  to 
local  excess  of  H-ions,  which  would  favor  autoh^sis.  Conversely, 
hypertrophy  is  observed  with  abundant  blood  supply  which  tends  to 
keep  the  reaction  of  the  tissues  so  low  in  Il-ions  that  autolysis  is  held 
at  a  niiiiinniiii. 

CLOUDY  SWELLING  ^^< 

The  characteristic  appearance  of  organs  the  seat  of  cloudy  swell- 
ing, which  is  frequently  likened  to  a  "scalded"  appearance,  sug- 
gests that  the  change  consists  in  a  coagulation  of  the  cell  proteins, 
which  idea  is  supported  by  the  similarity  of  the  microscopic  changes 
observed  in  the  cells  and  the  earliest  microscopic  changes  observed  in 
cells  after  heating  gently  to  about  their  maximum  thermal  point. 
On  the  other  hand,  the  granules  in  cloudy  swelling  are  generally  de- 
scribed as  being  soluble  in  dilute  acetic  acid  and  dilute  KOH,  which 
indicates  that  they  are  not  the  result  of  ordinary  heat  coagulation. 
If  we  bear  in  mind,  liowever,  that  cloudy  swelling  probably  does  not 
represent  one  single  change,  it  may  be  possible  to  arrive  at  some 
understanding  of  the  chemical  changes  that  occur  in  the  process. 
Albrecht  ^°  considers,  with  good  reason,  that  we  may  have  a  granular 
appearance  of  cells  which  is  simply  an  exaggeration  of  the  normal 
granular  structure,  and,  although  it  may  be  observed  in  tissues  mod- 
erately affected  by  toxins,  or  in  starvation,  or  in  transitory  anemia, 
the  change  is  still  to  be  looked  upon  as  little  more  than  physiological 
in  response  to  stimuli  and  overwork.  Such  a  ' '  cloudy  swelling ' '  may 
also  occur  in  cells  in.  the  beginning  of  autolysis,  or  simply  under  the 
influence  of  salt  solution.  If  the  injury  is  greater,  however,  as  in 
profound  sepsis,  or  extreme  local  anemia,  the  granules  become  coarser, 
less  soluble  in  acetic  acid  and  KOH,  and  droplets  resembling  "myelin'' 
make  their  appearance.  If  the  injury  is  still  more  severe,  true  coag- 
ulation of  the  granules  occurs,  and  they  become  insoluble,  the  fatty 
droplets  become  more  prominent,  and  the  cell  reaches  a  condition 
that  may  with  propriety  be  termed  necrosis  or  fatty  degeneration,  or 
both.  There  is  no  very  sharp  line  separating  necrosis  and  cloudy 
swelling,  especially  if  we  consider  only  the  changes  in  the  cytoplasm. 
In  the  earliest  stages  the  granules  are  jierhaps  due.  in  some  cases,  to 
simple  aggregation  of  the  colloids,  without  the  development  of  a  true 
coagulation,  and  so  the  granules  are  still  soluble.  Possibly  bacterial 
toxins  may  also  cause  soluble  precipitates,  but  this  does  not  appear 
to  have  been  established.  Halliburton  has  shown  that  temperatures 
that  iiuiy  be  reached  in   high  fevei's  can  cause  turl)idity  in  solutions 

2»fSloni<>ns,  Bull.  .Toliiis  Hopkins  llosp.,  1914    (25).  l!)."). 

2RKjour.  Biol.  Chcni..  lltlfi    (25),  2(51. 

20  Review  of  ^enoral  features  bv  Landstcinev,   Zic-^lcr's  T^cilr..   1!)0:?    iX]).  2'M . 

30  Verb.  Deut.  Patb.  Ccsoll.,  100.3   (6),  (>:{. 


CLOUDY  fiWEIJAM}  395 

of  cell  i)r()t(Miis,  and  luMice  heat  j^reeipitation  may  hv  partly  responsi- 
ble for  the  tui'hidity  of  eells  in  cloudy  swi'lling,  l)ut  it  is  d()ul)tful  if 
the  o-raimles  thus  formed  would  be  soluble  in  acetic  acid.  A  careful 
discussion  of  the  character  and  characteristics  of  this  process  is  given 
by  Hell,^""  who  concludes  that  the  term  cloudy  swelling  is  sound  only 
as  a  gross  description,  since  microscopically  the  cells  may  be  found  to 
show  albuminous  »>i-anules,  or  fatty  metamorpliosis  or  simple  edema. 
"When  present,  the  granules  are  of  unknown  nature — tiiey  are  not 
identical  with  Altmann's  granules,  although  Aschoff  and  Ernst  ^"^ 
both  consider  that  many  of  them  are  derived  from  the  mitochondria. 
An  enormous  number  of  granules  may  be  present  in  the  renal  cells 
without  demonstrable  impairment  of  function.'"''  They  may  disap- 
pear during  acute  infections,  and  they  bear  no  constant  relation  to 
fatty  changes. 

We  may  si)eak  with  more  assurance  concerning  the  swelling  of  the 
cell,  and  attribute  it  to  an  edema  of  the  cell  contents,  it  having  been 
shown  that  in  cloudy  swelling  the  water  content  of  the  organs  is  in- 
creased.^^ This  might  be  produced  by  a  rise  in  osmotic  pressure  due 
to  abnormally  rapid  splitting  of  proteins  with  incomplete  oxidation 
of  the  substances  formed,  which  results  in  formation  of  many  crys- 
talloid molecules  with  high  total  osmotic  pressure,  from  a  smaller  num- 
ber of  colloid  molecules  with  almost  no  osmotic  pressure.  It  has  fre- 
quently been  shown  that  the  cell-walls  do  not  lose  their  semipermea- 
ble character  until  the  death  of  the  cell  occurs ;  hence  in  cloudy  swell- 
ing water  diffuses  in  much  more  rapidly  than  the  crystalloids  can 
ditfuse  out,^-  causing  a  hydropic  swelling.  This  hypothesis  is  sup- 
ported by  the  observations  of  Cesaris  Demel,^^  who  found  that  by 
modifying  the  osmotic  conditions  of  the  cells,  particularly  epithelial 
cells,  he  could  closely  reproduce  many  of  the  characteristic  features 
of  parenchymatous  degeneration.  It  is  possible,  also,  that  too  high 
concentration  of  crystalloids  within  the  cells  may  be  a  factor  in  the 
precipitation  of  the  cell  colloids.  In  view  of  the  fact  that  in  the 
earliest  stages  of  autolysis  histologic  and  microscopic  changes  closely 
resembling  those  of  cloudy  swelling  are  pronounced,  and  that 
organs  the  seat  of  cloudy  swelling  notoriously  undergo  autolysis 
with  extreme  rapidity  after  death, ^^"^  we  may  also  consider  that  this 
process  is  possibly  in  part  responsible  for  the  change  of  ordinary  in- 
tra vitani  cloudy  sw^elling.  The  appearance  of  fine  granules  of  lipoid 
substance  ^^    (myelin   or  "protagon"    (?))    in   cells  during  autolysis 

30a  .Tour.  Amer.  Med.  Assoc,  1013    (61),  4.5.5. 

30b  Verb.  Dent.  Path.  Gesellsch.,  1914   (17),  43  and  103. 

'oc  Shannon,  Jour.  Lab.  Clin.  :\red.,  1916    (1),  541. 

31  Sc'hwenkenbecber  and  Tnpaki,  Arch.  exp.  Patli.  u.  Pbarm..  1906    (55).  203. 

32  See  introdiu'torv  oliapter  conoerninu  osmosis:   also  discussion  of  edema. 

33  Lo  Sperimentale.   1905:   Cent.  f.  Path.,   1905    (16),  613. 
33a  See  MedisTeceanu.  Jour.  Exp.  :\Ied..  1914   (19).  309. 

34  0rgler,  Virchow's  Arch.,  1904  (176),  413;  Hess  and  Saxl,  ibid..  1910  (202), 
149. 


396  RETROGRESSIVE    CTIANGES 

and  durinj;  cloudy  swelling  is  in  support  of  this  idea,  and  chemical 
analysis  of  organs  showing  cloudy  swelling  gives  definite  evidence 
of  autolytic  decomposition  of  the  proteins  and  an  increase  in  the  water 
content.^''  Presumably  this  increase  in  water  is  the  cause  of  the  low- 
ered specific  gravity  of  organs  exhibiting  parenchymatous  degener- 
ation.^°  Landsteiner,  through  his  studies  of  cloudy  swelling  in  human 
material,  also  came  to  the  conclusion  that  autolysis  is  an  important 
element  in  its  production. 

Martin  H.  Fischer  ^^  applies  the  principles  of  colloidal  chemistry 
to  the  problem,  and  concludes  that  the  changes  of  cloudy  swelling 
may  be  ascribed  to  acids  developed  in  the  cell.  Electro-negative 
proteins  in  the  cell  are  precipitated  by  weak  concentrations  of  acids, 
fonning  the  granules  in  the  cells,  which  can  be  dissolved  again  by  a 
stronger  concentration  of  acid  as  in  the  characteristic  clearing  of 
granular  cells  by  acetic  acid.  The  swelling  is  explainable  by  the 
increased  affinity  for  water  of  other  cell  proteins  under  the  influence 
of  acids.  This  theory  is  supported  by  good  experimental  evidence 
and  has  much  in  its  favor,  the  chief  question  being  whether  the 
blood  cannot,  under  ordinary  conditions  of  circulation,  furnish  suf- 
ficient neutralizing  salts  to  prevent  adequate  acidification  in  the  cells 
to  cause  cloudy  swelling. 

35  Verb.  Deut.  Path.  Gesell..  1903   (6).  76. 

36  See  Olsho,  Arch.  Int.  Med.,  1908  (2),  171. 

37  "Oedema  and  Nephritis,"  New  York,  1915,  p.  455;  also  Zeit.  Chem.  u.  Indust. 
Colloide,  1911    (8),  159. 


CHAPTER    XIV 

RETROGRESSIVE  CHANGES   (Continued) 

Fatty,  Amyloid,  Hyaline,  Colloid,  and  Glycogenic  Infiltration 
and  Degeneration 

FATTY  METAMORPHOSIS 

In  1847,  ill  the  tirst  number  of  his  Archiv,  Virchow  divided  the 
forms  of  fatty  changes  that  may  occur  in  pathological  conditions 
into  two  groups — ''infiltration"  and  "degeneration" — a  division 
tliat  has  since  become  classical.  By  infiltration  he  indicated  the  ex- 
cessive accumulation  of  fat  in  the  cells  in  the  form  of  large  drop- 
lets, without  destruction  of  the  nucleus  or  irreparable  damage  to  the 
cells,  and  by  the  use  of  the  term  infiltration  he  implied  his  belief 
that  the  fat  entered  the  cell  from  without.  When  the  fat  remained 
in  the  form  of  fine  droplets  and  the  cell  became  much  disintegrated, 
Virchow  considered  that  the  fat  was  derived  from  the  breaking 
dow^^  of  the  cell  proteins,  and  hence  the  process  was  considered  to 
be  a  fatty  degeneration  of  the  protoplasm.  Since  that  time  scarcely 
any  other  subject  in  pathology  has  been  more  warmly  discussed  than 
that  of  the  origin  of  the  fat  in  fatty  degeneration,  and  an  appalling 
amount  of  literature  has  accumulated  concerning  the  questions  in- 
volved. It  will  be  impossible  to  give  more  than  the  essential  facts 
that  have  been  developed,  referring  the  reader  for  the  full  details 
of  the  discussion  and  evidence  to  the  numerous  compilations  of  litera- 
ture, particularly  those  of  Rosenfeld,^  and  to  the  original  articles 
cited  in  the  text. 

PHYSIOLOGICAL  FORMATION  OF  FAT 

Concerning  the  normal  formation  of  fat  we  may  summarize  the 
evidence  as  follows : 

1  "Fat  Formation,"  Erfrelmisse  der  Physiol..  Al)t..  1,  1002  (1),  ti.")l:  ibid..  190.3 
(2),  50.  Also  see  discussion  in  the  Verh.  Dent.  Path.  Oosell..  1004  (Ti).  .37-108, 
and  the  review  bv  Leathes  in  his  ''Prolilems  in  Animal  ^Metabolism."  IflOO,  pp. 
71—121,  and  "Tlie  Fats,"  ^ionoGrraphs  on  Biochemistry,  London,  1010;  von  Fiirth, 
"Chemistry  of  Metabolism,"  Amer.  Transl..  New  York.  1916.  Concerning  theories 
of  role  of  li])ase  in  fat  metabolism  see  Chap.  iii.  Other  reviews  of  literature  on 
patholo£rical  fat  formation  bv  Christian.  .Johns  Hopkins  ITosp.  tiull..  IflOo  (16), 
1:  Lohlein,  Virchow's  Arch.,'  190.5  (180),  1;  Pratt.  Johns  TTopkins  IJosp.  Bull., 
1904  (15),  301  (particular  reference  to  heart)  ;  Wohlpemuth.  JTandbucli  d.  Bio-- 
chem.,  1909,  TIT  (1),  150;  :Ma?nus-Levv  and  Mever,  ihid..  1910,  JV  (1).  445; 
Dietrich,  Erpebnisse  der  Pathol.,  1909.  XIII  (2),  283.  Concerning  Obesity  see  v. 
Bercrmann,  Handbuch  d.  Biochera.,  1910.  IV  (2),  208.  Later  references  of  im- 
portance cited  in  the  text. 

397 


398  RETROGRESSIVE  CHANGES 

(1)  A  large  proportion  of  the  fat  of  the  body  comes  from  the 
fat  taken  in  the  food,  as  also  does  the  fat  of  the  milk.  This  can  be 
shown,  as  Rosenfeld  particularly  demonstrated,  by  starving-  an  ani- 
mal until  it  is  as  free  from  fat  as  possible,  then  feeding  with  a  large 
amount  of  some  fat  that  is  of  a  type  different  from  that  normally 
found  in  the  animal ;  the  new  fat  that  is  then  laid  up  in  the  fat  de- 
pots of  the  animal  will  partake  of  the  characters  of  the  fat  given  in 
the  food.  In  case  the  animal  is  lactating,  the  milk-fat  will  also  resem- 
ble the  fat  of  the  food.  As  a  matter  of  fact,  the  bod}-  fat  is  not  of 
constant  composition,  even  in  the  same  individual ;  it  varies  greatly 
with  age,  having  much  less  olein  in  infancy  than  in  later  years,  vary- 
ing somewhat  in  composition  in  the  different  fat  depots  in  the  same 
body,  and  apj^areiitly  being  more  or  less  modified  hy  diet. 

(2)  Fat  may  also  be  formed  from  carbohydrates.  According  to 
Rosenfeld,  this  fat  differs  from  the  fat  formed  on  mixed  diet  in  ha'v- 
ing  less  olein  in  proportion  to  the  palmitin  and  stearin,  and  it  is  de- 
posited particularly  in  the  subcutaneous  and  mesenteric  tissues  rather 
than  in  the  liver.  ]\Ian  does  not  seem  to  form  fat  readily  from  car- 
bohydrates, but  rather  burns  them  to  protect  his  proteins:  on  the 
other  hand,  swine  and  geese  readily  form  fat  from  carbohydrates. 
As  the  fatty  acid  radicals  of  ordinary  fat  (CigH„|,,Oo,  Cif.,H^2025 
CigHg^Oo),  are  much  larger  than  the  carbohydrate  radicals,  a  process 
of  synthesis  must  be  involved  in  the  formation  of,  fat  from  carbo- 
hydrates.- 

(3)  Proteins  are  a  possible  source  of  fat,  but  it  has  not  been  estab- 
lished that  they  are  either  a  common  or  an  important  source  of  fat  in 
either  physiological  or  pathological  conditions,  or,  indeed,  that  they 
really  ever  do  form  fat.  Upon  this  statement  rests  our  present 
tendency  to  refute  the  long-cherished  conception  of  fatty  degeneration 
as  a  true  degeneration  of  cell  proteins  into  fat,  as  suggested  by  A^ir- 
chow.  This  view  was  supported  by  the  earlier  work  of  Voit  and  his 
school,  who  believed  that  they  had  demonstrated  that  animals  could 
form  fat  from  protein  food,  and  their  work  was  for  a  long  time  ac- 
cepted as  correct.  Later  Pfliiger  and  his  pupils  pointed  out  what 
seem  to  have  been  essential  errors  in  these  investigations,  and,  after 
much  discussion  and  experimentation,  the  majority  of  physiologists 
now  support  the  view  advanced  in  the  sentence  opening  this  para- 
graph. Since  proteins  contain  carbohydrate  groups,  and  since  fats 
can  be  formed  from  carbohydrates,  the  ]iossibility  of  the  foi-mation  of 
fats  from  the  proteins  in  this  indirect  way  cannot  be  denied.  It  is 
also  possible  that  the  nitrogen-containing  groups  may  be  split  out  of 
the  amino-acids  of  the  protein  molecule,  and  that  the  non-nitrogenous 
residues  can  then  be  built  up  into  fatty  acid  molecules  as  large  as  the 

2'1'liis.  Ma^'inis  Levy  siifjposts,  may  1k'  iicc()iii|plislicil  (lirduiih  lactic  acid  wliicli 
is  forniod  from  siifjar.  and  tlicn,  after  reduction  to  an  aldciiydo.  several  of  tliese 
molecules  are  eombined  into  the  liijrlier  fatty  aeid.     See  Loathes,  loc.  cit.,  p.  S2. 


I'Al  II()I.(K1/<'M.  J-'AT  ACCVMILATION  399 

molecules  of  stearic,  palmitic,  and  oleic  acids;  l)ut  wc  have  no  proof 
that  either  of  tliese  i)roces.ses  occurs  in  the  normal  cell  or  in  the  cell 
that  is  undergoing  degeneration. 

PATHOLOGICAL  FAT  ACCUMULATION 

For  a  long  time  fatty  degeneration  was  looked  upon  as  one  of  the 
chief  evidences  that  fat  was  formed  directly  from  protein,  for  the 
cell  protoplasm  seemed,  morphologically,  to  be  changed  directly  into 
fat  in  this  process.  Additional  support  was  also  claimed  from  the 
sujijiosed  increase  in  fat  in  the  ripening  of  cheese ;  ^  from  the  forma- 
tion of  abundant  fat  by  maggots  living  in  fat-poor  blood  or  fibrin ; 
and  by  the  apparent  conversion  of  proteins  into  fatty  acids  and  soaps 
in  the  postmortem  change,  aclipocere.  But  it  has  now  been  well 
established  that  there  is  no  true  conversion  of  protein  into  fat  in 
the  fatty  degeneration  produced  experimentally  by  poisoning  with 
pho-sphonis,  etc.,*  and  the  other  supposed  instances  of  fat-formation 
above  cited  have  been  discredited  by  various  methods  which  it  will 
not  serve  our  purpose  to  discuss  here,  beyond  mentioning  that  one  of 
the  chief  sources  of  error  lies  in  the  fact  that  many  fungi  and  bac- 
teria ^  can  form  fat  from  protein. 

It  having  been  rendered  probable  that  fat  was  not  formed  by  dis- 
integration of  the  protein  of  the  degenerating  cells,  it  remained  to  de- 
termine what  the  source  of  the  fat  observed  in  the  cells  under  patho- 
logical conditions  might  be.  and  this  part  of  the  problem  has  been 
largely  cleared  up  by  Rosenfeld.  This  investigator  proceeded  as  fol- 
lows: Animals  were  starved  until  they  were  extremely  poor  in  fat, 
then  fed  upon  easily  identified  foreign  fats,  such  as  mutton  tallow 
(which  has  a  high  melting-point  and  can  combine  with  little  iodin) 
or  linseed  oil  (which  has  a  low  melting-point  and  can  combine  with. 
much  iodin).  The  animals  under  these  conditions  laid  up  in  their 
fat  depots,  including  the  liver  as  well  as  the  subcutaneous  tissues, 
large  quantities  of  these  foreign  fats.  By  starving  again  for  a  few 
days  the  foreign  fat  was  removed  from  the  liver,  leaving  still  a  large 
amount  in  the  other  storehouses,  and  the  animals  were  then  poisoned 
with  phosphorus  or  other  poisons  that  cause  a  typical  fatty  degener- 
ation of  the  liver  and  other  viscera.  When  the  fat  was  extracted  from 
the  fatty  liver  of  these  animals,  it  was  found  that  the  new  fat  that 
had  appeared  in  the  liver  during  the  process  was  not  normal  dog  fat 
C which  it  should  have  been  if  formed  by  degeneration  of  the  cell  pro- 
teins), but  was,  in  part,  of  the  same  type  as  the  foreign  fat  which 
the  animals  had  deposited  in  their  subcutaneous  tissues  and  other  fat 

3  Even  tlie  increase  of  fat  in  ripcninsr  cheese  is  doiilitful  (Xierenstein.  Proc. 
Royal  Soc.  B.,  1911    (S.3).  .301:  Kondo.  P.iochem.  Zeit..  1014   (50),  113). 

4  See  Tavlor,  Jour.  Exp.  :Med.,  1800  (4),  300;  Shihata,  Biochem.  Zeit.,  1011 
(37),  345. 

5  See  Beebe  and  Buxton,  Amer.  Jour,  of  Plivsiol.,  1005  (12),  466;  Slosse,  Arch. 
Internat.  Physiol.,  1004   (1),  348. 


400  RETROGRESSIVE  CHANGES 

storehouses.  Furthermore,  it  was  found  that  animals  stai*ved  to  an 
extremely  low  fat  content  do  not  develop  the  typical  fatty  liver  of 
phosphorus-poisoning:,  a  fact  which  Lebedeff  had  already  noted  in  a 
case  of  phosphorus-poisoning  in  an  emaciated  patient.  Of  similar 
sigiiiticance  is  the  fact  that  in  fatty  human  livers  the  iodin  number, 
normally  high,  falls  as  the  amount  of  fat  increases  until  it  is  ap- 
proximately that  of  adipose  connective  tissue.®  Therefore,  it  seems 
evident  that  the  fat  accumulating  in  the  liver  during  fatty  degenera- 
tion is  not  derived,  as  Virchow  thought,  through  a  transformation  of 
cell  proteins  into  fat,  hut  rather  is  an  infiltrated  fat  brought  in  the 
blood  from  the  fat  deposits  of  the  body  to  the  disintegrating  organ. 
This  work  has  since  been  corroborated  and  extended  by  many  ob- 
servers, and  its  correctness  can  now  hardly  be  questioned.'  "Fatty 
degeneration,"  therefore,  differs  from  "fatty  infiltration"  chietiy  in 
the  fact  that  in  the  former  the  process  is  associated  with  serious  in- 
jury to  the  cell,  caused  by  the  action  of  toxins  or  loss  of  nutrition, 
while  in  the  latter  the  cell  is  not  seriously  injured  and  is  capable  of 
returning  to  its  nonnal  condition  whenever  the  fat  is  removed.* 

Fatty  "Degeneration"  without  Infiltration. — By  showing  that 
new  fat  in  fatty  livers  is  infiltrated  fat,  Rosenfeld  did  not  entirely 
clear  up  the  subject,  for,  in  the  course  of  his  analyses  of  organs  that 
were  macro-  or  micro-scopically  the  seat  of  fatty  degeneration,  he 
found  that  there  is  not  always  any  correspondence  between  the  amount 
of  fat  that  seems  to  be  present,  as  determined  by  microscopic  methods, 
and  the  amount  that  chemical  analj^sis  shows  to  be  present.  This 
is  particularly  true  of  the  kidney.  Thus,  the  amount  of  fat  and 
lipoids,  or  lipins  (to  adopt  the  more  comprehensive  term  recom- 
mended by  Rosenbloom  and  Gies  ^  to  include  neutral  fats,  fatty  acids, 
soaps,  lipoids,  and  their  compounds),  present  in  normal  kidneys  (dog) 
was  found  to  vary  between  18.5  per  cent,  and  29.12  per  cent,  of  the 
dry  weight,  the  average  being  21.8  per  cent. ;  whereas,  after  producing 
a  typical  "fatty  degeneration"  by  means  of  phosphorus  and  other 
poisons,  the  lipin  content  was  still  found  to  be  between  16.9  per  cent, 
and  22.6  per  cent.^"     In  all  instances  the  amount  of  lipins  in  kidneys 

fi  Leathos,  Lanect,  Feb.  27,  1000;  Hartley  and  IMavrojiordato,  Jour.  Patli.  and 
Bact.,    1008    (12),  371;    Jaekson  and  Peare'e,  .Tour.   Kxp!  ^led..    1007    (0),   57S. 

7  Sehwalbe  (Verli.  der  Deut.  Path.  Cesell.,  1003  (0).  71)  claims  tliat  in  a  sim- 
ilar way  iodin  eomjiounds  of  fat  can  lie  demonstrated  to  lie  Iransjiorted  into  tlie 
fatt}"  organs.  Tlis  analyses  were  merely  qualitative,  and  hy  (|uantitative  deter- 
minations I  was  unable  to  corroborate  his  conclusions  (Zeit.  f.  plivsiol.  Chem., 
1905    (45),  412). 

8  A  strikinfT  proof  of  the  lack  of  injury  associated  Avitli  fatty  in(iltra(ion  is 
shown  by  the  fatty  infiltration  frequently  seen  in  the  liver,  especially  of  alcoholics, 
in  which  it  may  be  diOicult  to  find,  microscopically,  any  cell  cytojdasm  Ixn-ause 
of  the  fat,  the  tissue  looking  like  fatty  areolar  tissue;  and  yet  there  may  be  no 
clinical  evidence  wliatevcr  that  tlie  lix'cr  function  has  be(Mi  im])aire(I  i)y  tlie 
jirocess. 

!>  Bioclicm.   I'.iillctiii,    1011    (1),   -A. 

10  Concerning  the  normal  intracellular  fats  see  introductory  chapter. 


I'ATHOlJXilC.lL  FAT  ACri'Ml  I.ATIOS  401 

sliowiiiy  typiral  Tatty  degoueratiuu  under  the  microscope  was  found 
equal  to  or  less  than  the  normal  amount — it  was  never  increased. 
The  same  conditions  were  found  to  obtain  in  human  kidneys  that 
showed  fatty  nietaniorpliosis.  .Microscoj)ic  examination  of  s])ecimeiis 
stained  with  tlie  specific  fat  stains,"  therefore,  uives  no  indication  of 
tlie  amount  of  fat  contained  in  a  degenerated  kidney.  A  pathok)<iic 
kidney  containing:  16  per  cent,  of  lipins  (18  per  cent,  is  about  the  aver- 
age amount  in  noi-mal  human  kidneys)  inay  sliow  extreme  "fatty  de- 
generation" under  the  microscoi)e,  whereas  anotlier  kidney  may  con- 
tain as  mucli  as  23  per  cent,  of  lipins.  yet  not  show  any  fat  whatever 
by  staining  methods. 

The  explanation  of  tliis  remarkable  discrepancj'  is  as  follows: 
Every  tissue  and  organ  seems  to  contain  a  greater  or  less  amount  of 
lipins,  varying  from  5  per  cent,  to  20  per  cent,  of  the  total  diy  weight 
of  the  organ  in  the  case  of  most  of  the  important  tissues,  yet  this  is 
usually  held  in  such  a  form  that-  it  cannot  be  stained  by  any  stains 
available  for  the  purpose.  Thus  in  the  kidneys,  as  before  remarked, 
we  may  have  as  much  as  23  per  cent,  of  lipins  present  and  yet  be  unable 
to  stain  any  of  it  by  ordinary  methods.  The  greater  part  of  this  seems 
to  be  essential  to  the  cell,  for  it  cannot  be  removed  by  the  most  extreme 
starvation ;  e.  g.,  the  liver  of  the  most  emaciated  dogs  may  contain 
10  per  cent,  to  20  per  cent,  of  fatty  substances.  Furthermore,  the 
same  resistance  is  shown  by  part  of  the  fat  to  extraction  with  ether. 
A  certain  proportion  of  the  fat  can  be  extracted  readily  in  twenty- 
four  hours  or  less  by  ether,  but  after  this  time  no  more  can  be  made 
to  leave  the  tissues.  Apparently  the  rest  of  the  fat  is  held  in  a  com- 
bination (which  seems  to  be  chemical  rather  than  physical)  that  is 
insoluble  in  ether,  and  a  large  proportion  of  this  fixed  material  is  not 
simple  fat,  but  lecithin,  cholesterol,  and  compounds  of  these  lipoids. 
It  has  also  been  demonstrated  that  fatty  acids  can  combine  with 
amino-acids  to  form  compounds  (lipo-peptids)  very  similar  in  their 
properties  to  these  "masked"  fats.^-     B3'  digesting  the  tissue  for  a 

11  Fat-staining  involves  several  principles  of  interest  in  this  connection.  (See 
review  by  BuUard,  Jour.  Med.  Res.,  1012  (27),  55).  Osmic  acid  (OsOj),  the 
longest  used  for  tliis  puipose,  is  reduced  to  OsOo  by  oleic  acid,  impartintr  a  black 
or  dark-brown  color  to  the  fat:  but  it  does  not  stain  saturated  fatty  acids,  sucli 
as  palmitic  or  stearic  acid.  Thus,  Christian  found  in  pneumonic  exudates  fat 
that  stained  by  otlier  methods  but  not  by  osmic  acid,  apparently  l)ecause  it  con- 
tained no  oleic  acid  (Jour.  Med.  Research,  1003  (10),  100).  Sudan  III  and 
scarlet  R  {fat  poyvceau)  are  two  synthetic  dyes  Avhich  stain  fat  in  a  purely 
physical  way,  entering  and  remaining  in  the  fat-droplets  because  tliey  are  nuich 
more  soluble  in  fat  than  they  are  in  water  or  alcohol.  (Fully  discussed  by 
Michaelis  (who  introduced  scarlet  R)  in  Virchow's  Arch.,  1001  (164),  263;  and 
by  Mann,  "Physiological  Histology,"  p.  ."'06.)  These  stains  have  the  advantage 
of  staining  all  sorts  of  fats  and  not  staining  oilier  substances  that  may  reduce 
osmic  acid.  Fatty  acids  and  soaps  nmy  he  stained  witli  copper  acetate,  wliich 
forms  a  green  copper  salt,  and  thus  be  distinguislied  from  fats  ( Renda,  Yir- 
chow's  Arch.,  1000  (161),  104).  J.  Lorrain  Smith  (Jour.  Path,  and  Bact.,  1007 
(12),  1)  has  introduced  as  a  fat  dye  Nile  blue  sulphate,  which  forms  a  l)lue  salt 
with  free  fattv  acids,  while  neutral  fats  are  stained  red  bv  tlie  oxazone  base. 

12  Bondi,  Biochem.  Zeit.,  1900   (17),  .543. 

26 


402  RETROGRESSIVE  CHANGES 

short  time  by  pepsin,  however,  the  fixed  lipins  beeome  freed,  so  that 
they  can  then  be  readily  dissolved  out  in  ether.  AVe  see,  therefore,  that 
much  of  the  fat  of  normal  cells  is  so  firmly  combined  that  it  cannot 
be  dissolved  in  ether,  and  under  normal  conditions  all,  or  nearly  all, 
of  it  cannot  be  stained.  (This  applies  particularly  to  the  paren- 
chj'matous  org'ans;  the  fat  of  the  areolar  tissue  is  all  readily  ex- 
tracted— Taylor.)  By  the  use  of  Ciaccio's  method  for  microscopic 
demonstration  of  intracellular  lipoids.  Bell  ^^  has  been  able  to  dem- 
onstrate in  those  cells  that  are  fat-free  by  ordinary  methods  sufficient 
lipoidal  material  to  account  for  the  normal  "invisible  fat,"  which  is 
probabl}'  identical  with  the  "liposomes."  But  when  pathological 
changes  in  the  cells  result  in  decomposition  of  the  cell  protein  through 
autolysis,  or  produce  physical  clianges  in  the  colloids  that  hold  the 
lipins  emulsionized,  part  of  this  normally  invisible  fat  is  set  free,  and, 
becoming  visible,  "plianerosis,"  ^^  produces  the  so-called  "fatty  degen- 
eration." This  explains  the  observations  of  Rosenfeld,  cited  above, 
that  kidneys  may  show  much  fat  to  the  naked  eye  and  microscopically, 
when  they  actually  contain  even  less  than  normal  amounts  of  fat. 
Taylor  ^^  advanced  this  explanation,  and  supported  it  experimentally 
by  showing  that  during  fatty  degeneration  this  protected  fat  actually 
is  liberated,  some  two-thirds  becoming  ether-soluble  in  an  experiment 
performed  with  phosphorus-poisoned  frogs.  Mansfeld  ^^  also  found 
that  in  animals  poisoned  with  phosphorus,  the  proportion  of  fat  which 
is  present  in  a  form  free  from  protein  union  in  both  blood  and  viscera, 
is  increased,  while  the  firmly  bound  fat  is  decreased.  As  further 
support  may  be  mentioned  the  fact  that  organs  undergoing  experi- 
mental autolysis  show  microscopically  an  apparently  typical  fatty  de- 
generation, although  analyses  show  that  no  actual  increase  in  fat  oc- 
curs.'^ 

Relation  of  Anatomical  to  Chemical  Changes. — From  the  facts 
brought  out  in  these  various  experiments  we  must  consider  that  the 
anatomically  established  condition  of  "fatty  degeneration"  represents 
either  or  both  of  two  conditions :  ( 1 )  It  may  result  from  an  increase 
in  the  normal  quantity  of  fat  in  an  organ  undergoing  parenchymatous 
degeneration,  through  an  infiltration  of  fat  from  the  outside;  this 
is  particularly  true  of  the  fatty  degeneratioji  of  the  liver,  pre- 
sumably because  the  liver  normally  receives  the  relatively  saturated 
body  fats  to  work  them  over  into  the  more  labile  desaturated  fats;  (2) 

i3lntornat.  Monats.  Anat.  u.  Physiol.,  mil  (28),  297;  Jour.  :\I(m1.  I^cs..  1011 
(24),  5.39. 

14  Klompcrer,  Dent.  mod.  Wocli..  1909    (35),  89. 

in  .Tour.  Med.  Research,  190.3    (9),  59. 

1'!  Pfliipor's  Arch.,  1909   (129).  ti;\ 

17  Dietrich,  Arb.  path.  Inst.  Tiilrngeu,  1900  (5).  IT.  3;  Hess  and  Saxl.  Virchow's 
Arch.,  1910  (202),  149:  Ohta.  T?iochen).  Zeit..  1910  (29).  1;  Siiihata,  ihid..  1911 
(31),  321.  Tile  sif,'nificance  of  tlie  increase  of  li|iins  observed  in  ])erfused  l<idncvs 
by  dross  and  Vorpaiil  is  made  duulitful  1)V  ilie  urlirh^  of  riKhMliill  mid  lleiulrix.. 
Jour.  Biol.  Chem.,  1915   (22),  471. 


PATHOLOGICAL  FAT  ACCUMULATION  403 

or  there  may  be  no  increase  in  the  total  amount  of  fat,  but  the  invisi- 
ble fat  becomes  visible  through  autolysis  of  the  cell  proteins.  Thus, 
Bainbridge  and  Lcathes  '®  found  that  after  ligation  of  the  hepatic 
artery  there  is  a  marked  fatty  degeneration  of  the  liver,  without  an 
increase  in  the  amount  of  fat  according  to  analysis.  (3)  Finally,  of 
course,  both  factors  may  occur  together.  Of  these  various  forms,  in 
only  the  first  would  the  chemist  consider  the  organ  "fatty,"  although 
from  a  morphological  standpoint  the  second  form  is  entitled  to  rank 
as  a  true  "fatty  degeneration,"  and  the  form  that  will  occur  seems 
not  to  depend  upon  the  cause  of  the  cell  injury,  but  ratlier  upon  the 
organ  under  consideration.  In  a  study  of  the  relation  of  the  morpho- 
logical to  the  chemical  changes  Rosenfeld  ^^  arrived  at  the  following 
results : 

Normal  human  hearts  contain,  on  an  average,  15.4  per  cent,  of  lip- 
ins  ;  the  hearts  showing  fatty  degeneration  contain  20.7  per  cent,  on  an 
average.^""  The  pancreas,  which  normally  contains  15.8-17.4  per  cent., 
also  contains  an  increased  amount  when  showing  fatty  degeneration. 
The  liver,  however,  takes  on  by  far  the  greatest  amount  of  fat  after 
"steatogenetio"  poisons,-"  and  the  microscopic  picture  usually  gives  a 
very  good  approximation  of  the  amount  of  lipins  it  contains.^°^  Ap- 
parently in  these  organs  any  excessive  fat  above  the  normal  is  obsei'va- 
ble  microscopically,  although  the  normal  lipin  content  is  not,  and  only 
in  these  three  organs  could  Rosenfeld  find  an  actual  increase  in  fat 
after  poisoning-  with  phosphorus,  etc.  It  would  seem,  on  the  other 
hand,  that  there  is  not  often  a  real  increase  in  the  fat  content  of  the 
"fatty"  kidney.-^     Normal  spleen  contains  14.2  per  cent,  of  lipins, 

isBiochem.  Jour.,  1906   (2),  25. 

i9Berl.  klin.  Woch.,   1904    (41),  587. 

19a  The  amount  of  phospho-lipins  in  the  heart  is  usually  nearly  constant,  but 
alimentary  fat  may  accumulate  in  the  myocardium  under  certain  conditions. 
See  Wegelin,  Berl.'klin.  Woch.,  1913  (50),  2125;  Bullard,  Amer.  .Tour.  Anat., 
1916    (19),  1. 

20  In  fatty  livers  in  phosphorus-poisoning  the  amount  of  fat  may  reach  75  per 
cent,  of  the  dry  weight.  Accompanying  the  fat  increase  arc  increase  in,  water 
and  a  relative  or  absolute  decrease  in  proteins,  probably  due  to  cell  autolysis. 
In  acute  yellow  atrophy  a  similar  decrease  in  protein  occurs,  but  without  an  in- 
crease in  fat.      (See  v.  Starck,  Deut.  Arch.  klin.  Med.,  1884  (35),  481.) 

20a  See  Helly  (Beitr.  path.  Anat.,  1914  (60),  1)  who  examined  100  human 
livers  Avhich  showed  all  variations  in  microscopic  fat  content,  and  chemically 
from  7.36  to  74.43  per  cent,  of  lipins  (dry  weight).  He  found  tliat  there  was 
usually  a  good  correspondence  between  microscopic  appearance  and  analytic  re- 
sults, altho  some  marked  and  unexplained  discrepancies  were  observed.  Gener- 
ally the  fat  content  was  from  10  to  30  per  cent,  of  the  dry  weight,  with  19  to 
21  per  cent,  the  most  common  figures.  When  there  is  much  fat  present  in  the 
liver  the  fat  content  of  the  bile  is  increased  (Le  Count  and  Long,  Jour.  Exp. 
Med..  1914    (19),  234). 

21  This  is  contradicted  by  Landsteiner  and  ^lucha  (Cent.  f.  Path.,  1904  (15),. 
752)  and  by  Lohlein  (Virchow's  Arch.,  1905  (180),  1)  and  Bosenthal  (Deut. 
Arch.  klin.  INIed.,  1903  (78),  94),  but  is  supported  bv  Orgler  {ibid.,  1904  (176), 
413),  and  Dietrich,  Verb.  Deut.  Path.  Gesell.,  1907  (11),  10.  See  also  the  later 
studies  by  Rosenfeld  on  the  effects  of  various  steatoffenic  poisons  on  different 
organs,  in  Arch.  f.  Exp.  Path.  u.  Pharm.,  1906  (55),  179  and  344.  It  is  probable 
that  the  truth  lies  between  tlie  opposing  views,  namely,  the  kidney  may   under 


404  RETROGRESSIVE  CHANGES 

and  lung  17.3  per  cent.,  but  in  both,  "fatty  degeneration"  results  in  a 
lowering  of  this  quantity.  Degenerations  in  the  nervous  tissue,  which 
Yirchow  considered  the  best  evidence  of  the  conversion  of  protoplasm 
into  fat,  also  show  a  marked  decrease  in  lipins,  and  voluntary  muscle 
shows  no  increase  in  the  normal  quantity  after  poisoning.  In  general, 
these  experiments  support  the  contention  of  Taylor  concerning  the 
disclosure  of  the  invisible  fat  through  autolysis.--  An  explanation  of 
many  of  the  discrepancies  lies  in  the  newer  studies  on 

The  Relation  of  the  Lipoids  to  Fatty  Metamorphosis.-^ — Until  within 
a  few  years  the  significance  of  the  intracellular  ]ii)oids  in  fatty  degen- 
eration and  related  processes  was  not  appreciated,  beyond  the  fact 
that  in  most  organs  showing  fatty  changes  the  quantity  of  cholesterol 
and  lecithin  is  not  greatly  changed.  In  1902  Kaiserling  and  Orgler 
described  under  the  non-committal  name  of  "myelin"  certain  intra- 
cellular droplets  that  may  be  found  in  the  cells  of  the  normal  adrenal 
cortex,  and  in  amyloid  kidnej^s,  pneumonic  exudates,  tumor  cells, 
retrogressive  thymus  tissue,  corpus  luteum,  and  bronchial  secretions, 
and  which  differ  from  fat  in  being  doubly  refractile  (anisotropic) 
when  viewed  through  Nicoll  prisms,  and  in  staining  but  slightly  gray 
with  osmic  acid,  although  taking  up  other  fat  stains  well. 

As  explained  in  Chapter  i,  the  myelins  are  probably  mixtures  of 
lipins,  cholesterol-esters  being  prominent,  and  in  many  conditions  in 
which  fat-like  vacuoles  are  prominent  in  cells,  leading  to  the  diagnosis 
of  fatty  degeneration,  these  substances  are  responsible,  presumably 
having  been  liberated  from  combination  with  the  cell  proteins  in  some 
cases,  in  others  actually  being  increased  in  the  cell.  This  condition, 
which  Aschoff  refers  to  as  a  cholesterol-ester  fatty  metamorphosis,  is 
especially  seen  in  the  parenchyma  cells  derived  from  the  urogenital 
anlage — that  is,  the  adrenal  cortex,  kidney,  testicle  and  corpus  lu- 
teum. Aschoff  states  that  doubly  refractile  droplets  can  be  formed 
by  lecithin  and  phosphatids  generally,  oleates,  cholesterol  esters,  cho- 
lesterol when  dissolved  in  phosphatids  or  fats  or  fatty  acids,  as  well 
as  by  cholesterol  esters  dissolved  in  fats.  Of  these  the  most  im- 
portant quantitatively  is  the  cholesterol  ester  group,-*  and  the  anal- 
yses of  Windaus  have  sliown  that  in  ])ath()l()gical  processes  the  increase 

some  conditions  take  up  fat  from  the  blood,  but  it  does  so  to  a  much  loss  oxtont 
than  tlie  liver,  and  it  may  sometimes  show  marked  fatty  change  anatomically 
without  correspondin<j  increase  chemically. 

--  I'ieces  of  tissue  im])lant('d  into  animals  may  show  a  peripheral  fatty  meta- 
morphosis or  intiltrat  ion,  xct  show  >i])on  analysis  a  decreased  fat  content 
(Dietrieh,  Verh.  Deut.  Path."  CJesellsch.,  IDOf)    (n),'212). 

-s  Literature  by  I.ieathea,  "The  Fata,"  London,  1!M0;  l^anp,  l\r<febnisse  der 
Physiol.,  1009  (8),  4()3,  also,  "Chemio  u.  Biochem.  d.  Lipoide,"  Berfrmann.  Wies- 
l)aden,  1011;  Kawamura,  Virchow'a  Arch.,  1012  (207),  400,  also  "Die  C'holester- 
inesteryerfettunfr,"  Fischer,  Jena,  1011;  AschofT,  Zie<rler's  Beitr.,  1000  (47),  1, 
also  Festschr.  f.  Unna,  1011,  p.  23;  Schultz,  Erpebnisse  d.  Pathol.,  1000  (XTITJ, 
253;  llanes,  Bull.  Johns  Hopkins  llosp.,  1012  (23),  77:  Anits.-hkow  and  Chala- 
tow.  Cent.  f.  Pathol.,  1013    (24),  1. 

n^See  also  Verse,  Ziegler's  Beitr.,  1011    (,'52),  1. 


I'AI  llOlAXiKWL  FAT  ACCUMULATION  405 

is  niuc'li  jiTeatiT  in  the  cholesterol  esters  than  in  the  free  cholesterol. 
Cholesterol  compounds  stain  differently  from  neutral  fats,  beino;  more 
yellow  than  red  with  Sudan  III,  and  iirayish  rather  than  black  with 
osinie  acid.  Path()l()j.iically  the  anisotropic  drojjlets  are  also  found 
especially  in  the  above-named  tissues,  but  also  in  tissues  the  site  of 
chronic  inflammation,  including:  the  mucosa  of  the  gall  bladder  where 
they  may  be  of  importance  in  the  formation  of  cholesterol  concre- 
tions. They  are  also  f(mnd  in  tlie  alveolar  epithelium  in  pulmonary 
intianmiation,  in  atheromatous  patches  in  arteries,  in  many  tumors, 
in  most  cells,"^  and  occasionally  in  varied  patholog-ical  tissues.-^'^  Per- 
haps the  most  conspicuous  deposits  are  in  the  epithelium  of  the  ''large 
white  kidneys,"  and  in  xanthomas.  In  Gaucher 's  disease  there  is  also 
a  remarkable  lipoid  accumulation  in  the  foamy  phagocytic  cells. -^'' 
According  to  ]\[unk  -"  true  lipoid  degeneration  always  means  a  serious 
injury  to  the  cell,  but  there  seem  to  be  many  exceptions  to  this.  In- 
deed, according  to  Anitschkow  and  Chalatow  {loc.  cit.)  the  feeding  of 
foods  rich  in  cholesterol  may  cause  the  appearance  in  the  liver  of  great 
quantities  of  anisotropic  droplets,  lipoid  deposits  in  the  aorta,  enlarge- 
ment of  the  adrenal  cortex,  and  the  presence  in  practically  all  tissues 
of  semifluid,  doubly  refracting  crystalline  structures  {cholesterol 
steatosis)  .'^^'^ 

In  cells  undergoing  autolysis  the  fat-like  "myelin"  droplets  which 
appear,  differ  from  the  above  in  not  being  anisotropic,  but  are  un- 
doubtedly closely  related  to  them  in  composition.  These  "myelin" 
droplets  are  also  found  in  cells  showing  cloudy  swelling,  presumably 
representing  cell  lipoids  set  free  through  changes  in  the  cell  proteins. 
They  are  characterized  by  staining  with  osmic  acid  but  not  by  sudan 
III,  which  shows  them  not  to  be  simple  fats  nor  yet  lipoids,  but  they 
are  undoubtedly  precursors  of  true  fatty  degeneration ;  "  they  prob- 
ably consist  chiefly  of  lecithin,  with  more  or  less  free  fatty  acids  and 
relatively  little  cholesterol    (Aschoff). 

It  is  possible  to  distinguish  the  lipoids  of  cells,  whether  normal  or 
pathological,  from  the  neutral  fats  by  means  of  Ciaccio's  method.-* 
This  consists  in  a  preliminary  treatment  with  bichromate,  which  ren- 
ders the  lipoids  insoluble ;  the  tissues  can  then  be  hardened  and  im- 
bedded by  the  usual  methods  which  remove  the  unchromated  fats, 
leaving  the  lipoids  stainable  by  Sudan  III.     By  this  method  Bell  -^ 

25Ciaccio,  Cent.  f.  Path.,  1013    (24),  .'50. 

25a  Patholotrical  deereape  in  lipoids  may  also  ho  ohscrved,  especially  in  the  adrenal 
cortex,  usnally  under  the  influence  of  toxic  ajients;  e.  g.,  Hirsch  found  a  marked 
decrease  in  delirium  tremens   (Jour.  Amer.  Med.  Assoc.,  1014   (fi.S),  2186). 

25b  See  Wahl  and  Richardson.  Arch.  Int.  Med.,  1016   (17),  2.38. 

20Virchow's  Arch..  1008    (104),  527. 

2CaSee  also  Anitschkow.  Dent.  med.  Woch.,  101.3  (30),  741;  ^Yosselkin.  Vir- 
ehow's  Arch.,  1013    (212).  22-5. 

27  Hess  and  Saxl,  Virchow's  Arch.,  1010    (202),  140. 

28  Cent.  f.  Path..  1000   (20),  771;  Arch.  f.  Zellf.,  1010   (5),  235. 

29  .Jour.  Med.  Pes..  1011    (24),  530. 


406  RETROGRESSIVE  CHANGES 

has  been  able  to  stain  the  lipoids  in  the  normal  kidney  and  other  tis- 
sues, in  sufficient  amount  to  account  for  all  the  so-called  "masked 
fat,"  which  thus  seems  to  be,  as  also  indicated  by  chemical  evidence, 
largely  lipoidal. 

Jastrowitz  -^'^  has  studied  the  relation  of  lipoids  to  fats  in  the  fatty 
changes  produced  by  various  means,  and  finds  that  in  severe  fatty 
changes  with  much  transported  fats  there  may  be  little  change  in  the 
lipoids;  with  blood  poisons  which  cause  little  increase  in  total  fats, 
the  lipoid  content  of  both  blood  and  organs  may  be  high ;  usually  the 
phosphatid  content  is  unchanged  or  slightly  increased,  but  it  may  be 
decreased.  The  proportion  of  cholesterol  to  neutral  fats  is  usually 
within  normal  limits  in  tissues  showing  fatty  changes.-*^  The  mito- 
chondria seem  to  be  compounds  of  phospholipins  with  proteins,  and 
these  agglutinate  and  form  fatlike  droplets  in  phosphorus  poison- 
ing ;  -^'^  presumably  they  play  an  important  role  in  fatty  metamor- 
phosis. 

Summary. — We  must  conclude,  therefore,  that  fatty  degeneration 
of  an  organ  means,  in  the  case  of  the  liver,  myocardium,  and  pan- 
creas, an  infiltration  of  neutral  fat  from  outside  into  cells  which  have 
been  degenerated  by  the  action  of  poisons  or  other  injurious  influ- 
ences, plus  a  certain  amount  of  apparent  increase  in  fat  because  of  the 
setting  free  of  previously  invisible  fats  and  lipoids  normally  present 
in  the  affected  cells.  In  the  kidney,  spleen,  and  muscles  an  increase 
of  fat  seldom  occurs  from  these  causes,  but  the  cells  may  show  a 
marked  fatty  metamorphosis  through  the  setting  free  of  the  invisible 
intracellular  fat  and  lipoids  by  autolytic  or  physico-chemical  changes. 
In  the  adrenal,  kidney,  and  often  in  other  tissues,  the  fatty  material 
present  in  the  cells  is  characterized  by  being  doubly  refractile,  and 
then  consists  chiefly  of  cholesterol  esters,  together  with  greater  or  less 
quantities  of  phosphatids,  fatty  acids,  soaps  and  neutral  fats. 

CAUSES  OF  FATTY  METAMORPHOSIS 

Nevertheless,  the  old  anatomical  distinction  of  infiltration  and  de- 
generation still  remains,  provided  we  do  not  hold  to  the  original  idea 
that  the  term  degeneration  implies  that  the  cell  protein  has  been  eon- 
verted  into  fat ;  for  we  must  recognize  that  under  some  conditions  the 
cells  may  take  up  great  quantities  of  fat  without  suffering  any  appre- 
ciable degenerative  changes,  whereas  in  other  instances  the  appear- 
ance of  fat  is  associated  with  marked  and  complete  disintegration  of 
both  nucleus  and  cytoplasm.  Furthermore,  we  have  yet  to  explain 
why,  under  some  conditions,  the  fat  is  removed  from  the  fat  depots 
to  be  stored  up  in  the  liver  or  other  organs.  By  appl.ving  the  com- 
monly accepted  ideas  concerning  fat  metabolism,  a  satisfactory  ex- 

28aZeit.  exp.  Palli.  u.  Tlior.,  1914   (15),  110. 
28bCzyhlarz  and  Fudis,  Biochem.  Zeit.,  ]!)14   (63),  1.31. 
28c  Scott,  Amer.  Jour.  Anat.,  1916    (20),  237. 


CAUSES  bl-  FATTY  M iriA  MOh'l'HOSIS  407 

planation  seems  to  be  possible.  Fat  is  always  utilized  and  trans- 
ported in  the  form  of  its  two  constituents,  fatty  acid  (or  soaps)  and 
glycerol,  which  are  diffusible  and  soluble.  It  enters  and  leaves  the 
cells  in  this  condition,  being  split  or  combined,  as  may  be  necessary  to 
produce  equilibrium,  by  the  action  of  lipase,  which  is  present  within 
the  cells  and  in  the  blood  and  lymph.  Under  normal  conditions  there 
is  little  free  visible  fat  in  the  cells  of  the  parenchymatous  organs, 
because  it  is  largely  used  up  througli  oxidation  of  the  glycerol  and 
fatty  acids  by  the  action  of  the  intracellular  oxidases.  AVhere  there 
is  abundant  lipase  and  but  little  oxidative  activity,  as  is  the  case  in 
the  areolar  fat  tissue,  fat  accumulates  in  large  amounts.  AVhen,  for 
any  reason,  the  oxidative  power  of  the  parenchymatous  organs  is  re- 
duced, fat  accumulates  in  them  as  it  does  in  the  fat  depots  normally, 
and  we  have  an  excess  of  fat  in  the  parenchymatous  cells ;  thus,  in 
pulmonary  tuberculosis,  severe  or  protracted  anemias,  etc.,  a  great 
accumulation  of  fat  occurs,  particularly  in  the  liver,  where  normally 
active  oxidative  processes  continually  balance  the  action  of  the  abun- 
dant lipase  of  the  liver-cells.  The  liver  being  normally  concerned  in 
the  preparation  of  fat  for  metabolism,  it  is  also  perfectly  possible  to 
have  an  accumulation  of  fat  in  the  nonnal  liver  merely  as  a  result  of 
increased  function,  and  hence  fatty  changes  may  be  purely  physio- 
logical in  this  organ. "^^ 

If  the  fat  accumulates  in  cells  that  are  structurally  normal  or 
nearly  so,  the  fat-droplets  fuse  together  under  the  pressure  of  the 
c\i:oplasm,  and  w^e  get  the  picture  of  a  typical  fatty  infiltration;  in- 
deed, the  only  tissues  in  which  we  get  this  typical  infiltration  are  the 
liver  and  the  fatty  areolar  tissue,  in  both  of  which  the  process  is  pre- 
sumably physiological  in  character  even  if  not  always  physiological 
in  degree.  If  the  cells  are  much  disintegrated  through  the  action  of 
the  poison, — e.  g.,  phosphorus,  bacterial  toxins,  etc., — the  accumulat- 
ing fat-droplets  are  not  crowded  into  one  large  droplet,  but  lie  free 
in  the  granular  debris  of  the  disintegrating  cell,  constituting  the  typi- 
cal appearance  of  fatty  degeneration.  Fatty  degeneration  is  usually 
brought  about  by  poisons,  while  abnormal  fatty  infiltration  depends 
usually  upon  decreased  oxidation,  due  to  lack  of  either  oxygen  or 
hemoglobin  in  the  blood.  If  the  anemia  is  extreme,  however,  the  cells 
degenerate,  and  then  we  find  a  true  fatty  degeneration  caused  by  lack 
of  oxygen.^"  Thus,  in  an  anemic  infarct  fat  accumulates  about  the 
periphery  of  the  dead  area,^^  probably  because  fatty  acids  and  glycerol 
diffuse  in  slowly  from  the  surrounding  parts  where  circulation  still 

29a  See  Coope  and  Mottram,  Jour,  of  Phvsiol.,  1914  (40),  23;  Ilellv,  Beitr.  path. 
Anat...  1914    (00),  1. 

"io  Mohr  (Zeit.  exp.  Path.,  1906  (2),  434)  denies  that  oxidation  is  decreased  in 
anemia;  and  in  a  man  with  but  about  half  the  normal  lunw  area  the  metabolism 
was  not  found  altered  to  anv  extent  bv  Carpenter  and  Benedict,  Amer.  Jour. 
Phvsiol.,  1909    (23),  412. 

siFischler,  Cent.  f.  Path.,  1902   (13),  417. 


408  h'KTh'OflHESSnE  CHANGES 

goes  on,  and  are  built  up  into  fat  by  tlie  cell  lipase,  for  in  anemic  areas 
the  intracellular  oxidases  cannot  destroy  these  substances  as  they  nor- 
mally do,  because  of  lack  of  oxygen.  The  accumulation  of  fat  in  dead 
areas  depends,  therefore,  on  the  fact  that  the  constituents  of  fat  can 
diffuse  into  the  dead  tissue,  whereas  the  oxyg-en,  being  held  in  the  cor- 
puscles, cannot  enter  the  anemic  area.^-  It  is  also  possible  that  where 
fat  is  set  free  by  autolysis  of  dead  tissue,  or  when  for  any  cause  free 
fat  or  lipoid  material  is  present  in  the  vicinity  of  living  cells,  it  may 
be  phag:ocyted  or  in  some  way  infiltrate  the  cells,  causing  a  fatty  meta- 
morphosis by  absorption  (Dietrich). 

It  is  to  be  supposed  that  poisons  also  cause  fatty  degeneration  in 
a  similar  way — by  interfering  with  oxidation.  We  have  much  evi- 
dence that  in  phosphorus,  chloroform,  and  other  poisoning  associated 
with  fatty  degeneration  of  the  liver,  oxidation  is  impaired.^^  If  we 
imagine  for  a  moment,  a  cell  in  which  oxidation  is  checked  by  any 
means,  we  shall  have  in  this  cell  the  lipase  and  the  proteolytic  en- 
zymes not  balanced,  as  they  normally  are  by  the  action  of  the  oxi- 
dases, and  hence  the  processes  of  cell  autolysis  and  of  the  accumula- 
tion of  fat  by  the  lipase  will  go  on  uncontrolled.  The  result  will  be 
a  disintegrated  cell  containing-  many  fat-droplets,  i.  e.,  fatty  degen- 
eration.^* In  cloudy  swelling  there  also  appear  droplets  stained  with 
osmic  acid  but  not  by  sudan  III,  which  Hess  and  Saxl  ^^  have  shown 
to  result  from  intravitam  cell  autolysis,  and  to  be  a  precursor  of  true 
fatty  degreneration. 

Work  with  cells  in  tissue  cultures  indicates  that  fatty  changes  of 
all  types  may  occur  independently  of  the  circulation.  Lambert  ^^"^ 
states  that  the  amount  of  fat  in  the  culture  cells  is  roughly  propor- 
tional to  the  amount  in  the  culture  medium,  and  cells  rich  in  fat  may 
move  actively  and  undergo  normal  mitosis.  Lewis,  however,  observed 
fatty  changes  in  cells  growing  in  fat-free  media,  and  made  the  espe- 
cially interesting  observation  that  cells  grown  in  2.5-3  per  cent,  alco- 
liol  will  show  a  rich  fat  accumulation.     Also,  an  accumulation  of  fats 

32  See  Griesser,  Ziegler's  Beitr.,  mil    (51),  11.5. 

■■^3  See  Welsch,  Arch.  int.  de  pliarm.  et  therap.,  1905    (14).  211. 

•"•*  Interference  with  oxidation  does  not  necessarily  imply  destruction  of  the 
oxidases.  As  yet  we  know  practically  notliinn;  concerninsi  the  oxidases  of  the 
cells  in  disease,  and  the  above  hyjiothesis  has  yet  to  be  demojistrated.  Duccheschi 
and  Almafjia  (Arch.  Ttal.  Biol.,  100.3  (.SO),  20)  found  the  normal  amount  of 
lipase  in  phosphorus-livers,  but  also  observed  no  decrease  in  ability  to  oxidixo 
salicylic  aldehyde,  which,  however,  ''ocs  not  prove  a  normal  ])ower  io  oxidize  fats. 
Gierke's  observation  (Zieji-ler's  Bcif'.,  1005  (37),  502)  tliat  "rlycoiren  and  fat 
accumulate  imdcr  identical  conditions  mifiht  be  cit(>d  as  judical injr  decreased  oxi- 
dative power.  Wells  (.Tour.  Exper.  ^Med.,  1010  (12),  007)  found  that  the  power 
of  liver  tissue  to  oxidize  i)urines  was  Tiot  decreased  bv  the  maximum  dearee  of 
fatty  defjeneration,  but  Waldvo<rel  (Deut.  Arch.  klin.  ]\Ied.,  1007  (SO),  :?42)  found 
that  obese  persons  can  burn  fatty  acids  which  arise  in  metabolism  less  readily 
than  normal;  and  Quinan  (Jour.  Med.  Bes.,  1015  (.32),  73)  found  the  ester- 
splittinp  lipolytic  enzymes  of  tlie  liver  much  reduced  in  the  liver  of  chloroform 
necrosis,  but  the  relation  of  these  esterases  to  true  lipases  is  Tiot  known. 

3n  Virchow's  Arch.,  1010    (202),  140. 

35n  Trans.  Assoc.  Amer.   I'hys.,  1013    (0),  03;  Jour.  Exp.  Afcd..  1014    (10).  30S. 


CAUSES  OF  FATTY   MET  A  MOI!  I'IKtsIS  409 

and  li])<)i(ls  in  i-clls  <:ro\vii  in  tlie  presence  of  sudi  steatogenetie  poi- 
sons as  pliosphorns  and  Oleum  pulegii  has  been  observed  by  others,'*"''' 
which  indicates  that  free  cells  behave  the  same  under  the  influence 
of  such  poisons  as  the  cells  of  the  fixed  tissues. 

Tlie  jirocess  of  nninaskin<|-  the  masked  fats  is  explained  by  M.  H. 
Fischer^'"'  on  a  pliysical  l)asis,  as  follows:  The  fats  of  the  cells  are 
distributed  as  an  emulsion  in  a  h3'dration  compound  of  water  with 
hydroi)hilic  colloids,  notably  proteins  and  soaps.  Such  an  emulsion 
breaks  down  whenever  the  hydropliilic  colloid  is  either  dehydrated  or 
diluted  beyond  certain  ranges.  As  the  usual  conditions  that  cause 
fatty  degeneration,  such  as  poisoning  with  phosphorus,  arsenic,  etc., 
or  local  circulatory  disturbances  with  local  acidosis,  all  tend  to  de- 
hydrate some  of  the  cell  colloids  and  to  dilute  others,  it  would  seem 
probable  that  the  appearance  of  the  fat  droplets  in  the  cells  is  the 
result  of  such  changes  in  the  colloids  that  previously  held  them  in  an 
emvilsion  too  fine  to  exhibit  readily  visible  fat  particles.  The  relation 
of  cloudy  swelling  to  fatty  degeneration  is  readily  explained  on  this 
basis,  as  follows:  When  a  local  acid  intoxication  of  a  cell  occurs, 
some  of  the  proteins  will  swell  and  others  will  precipitate,  resulting 
respectively  in  the  swelling  and  cloudiness  of  the  cells  characteristic 
of  cloudy  swelling;  but  at  the  same  time  the  emulsifying  capacity  of 
these  proteins  will  be  impaired,  penuitting  the  coalescence  of  the  fat 
droplets  and  the  resulting  picture  will  be  that  of  fatty  degeneration. 

Summary. — Fatty  metamorphosis  involves  changes  of  two  kinds. 
First,  infiltration  of  fat,  which  occurs  when  the  oxidative  power  of 
the  cells  is  decreased,  so  that  fat  is  not  destroyed,  but  is  accumulated 
from  the  blood  under  the  influence  of  the  lipase  of  the  cells;  if  there 
is  not  any  serious  injury  to  the  cells,  the  histological  changes  consist 
in  the  accumulation  of  one  or  a  few  large  droplets  of  fat  in  each  cell, 
constituting  the  condition  known  anatomically  as  "fatty  infiltration." 
This  occui*s,  pathologically,  chiefly  in  the  liver.  If  at  the  same  time 
the  cytoplasm  is  disintegrated  through  autolytic  changes,  the  fat- 
droplets  do  not  fuse,  but  remain  as  small,  more  or  less  discrete,  fat 
granules  among  the  granules  of  cell  debris,  constituting  the  micro- 
scopic picture  of  "fatty  degeneration";  this  condition  occurs  partic- 
ularly in  the  heart  and  liver. 

Second,  each  cell  contains  a  large  amount  of  fat  and  lipoids  (5-25 
per  cent,  of  its  dry  weight),  which  is  so  combined  that  it  cannot  be 
detected  microscopically ;  this  may  be  liberated  during  the  autolytic 
processes  and  colloidal  changes  of  cell  disintegration  and  become 
visible,  constituting  a  macroscopical  and  microscopical  degeneration, 
but  without  any  actual  increase  in  fat — this  condition  occurs  partic- 
ularly in  the  kidney  and  nervous  system.     Third,  a  combination  of 

35b  Krontowski  and  Poteff,  Beitr.  path.  Anat.,  1914   (58),  407. 
35C  Fischer  and  Hooker,  Science,  1916  (43),  468;  Fischer,  Fats  and  Fatty  Degen- 
eration, Wiley,  New  York,  191". 


410  RETROGRESSIVE  CHANGES 

both  of  the  above  processes — infiltration  of  fat  and  liberation  of 
masked  intraeelhilar  fat — may  occur  simultaneously  in  an  organ.^" 
Fourth,  in  certain  cells,  especially  in  the  kidney,  adrenal,  ovary  and 
some  tumors,  there  maj^  be  a  great  increase  in  the  lipoids  of  the  cell, 
^'lipoidal  degeneration,"  and  especially  of  cholesterol  esters  and  free 
cholesterol,  part  of  which  is  infiltrated  and  part  set  free  from  com- 
bination in  the  cytoplasm. 

PROCESSES  RELATED  TO  FATTY  METAMORPHOSIS 

ADIPOCERE 

This  apparent  transformation  of  the  substance  of  dead  bodies  into 
a  wax-like  material  was  for  a  long  time  looked  upon  as  evidence  of  a 
transformation  of  protein  into  fat,  but  in  the  light  of  more  recent  in- 
vestigations this  view  can  hardly  be  held.  Adipocere  is  the  product 
of  a  process  that  occurs  particularly  in  bodies  buried  in  very  wet 
places  or  lying  in  water,  and  results  in  an  apparent  replacement  of 
the  muscles  and  other  soft  parts  (but  not  the  glandular  organs)  by  a 
mass  consisting  of  a  mixture  of  fatty  acids  in  crystalline  and  amor- 
phous form,  and  soaps,  particularly  ammonium,  magnesium,  and  cal- 
cium salts  of  palmitic  and  stearic  acid  (the  oleic  acid  largely  disap- 
pearing during  the  process).  Analysis  of  a  sample  of  adipocere  (de- 
rived from  a  buried  hog),  by  Ruttan  and  Marshall  ^'^'^  showed  but  4.4 
per  cent,  of  calcium  soaps,  as  contrasted  with  67.5  per  cent,  of  palmitic, 
3.3  per  cent,  stearic,  5.24  oleic,  and  15.8  per  cent,  hydroxy  stearic 
acids;  94.1  per  cent,  of  the  adipocere  was  soluble  in  ether.  Ammonium 
and  other  soluble  soaps  were  absent.  The  hydroxy-stearic  acids,  which 
are  so  characteristic  of  adipocere,  are  formed  from  the  oleic  acid  of 
the  original  triolein.     But  0.18  per  cent,  of  nitrogen  was  present. 

The  resulting  material  is  absolutely  resistant  to  putrefaction,  and 
hence  remains  intact  for  many  years.  This  replacement  of  the  soft 
parts  is,  however,  only  apparent,  for  the  total  weight  of  a  body  in 
this  condition  is  much  lighter  than  that  of  the  original  body ;  indeed, 
one  is  always  surprised  at  the  light  weight  on  lifting  such  a  specimen. 
Adipocere  occurs  almost  exclusively  in  fat  bodies,  and  it  seems  probable 
that  all  the  soaps  and  fatty  acids  found  are  formed  from  the  orujinal 
fats  of  the  corpse.^^^  These  gradually  flow  into  the  places  left  by  the 
disintegrating  muscle,  etc.,  a  process  that  occurs  readily  in  cadavers, 
according  to  Zillner;  ^^  or  the  infiltration  may  be  accomplished  through 

38  The  above  conception  of  the  processes  involved  in  fatty  metamorphosis  is 
more  fully  discussed  Ijy  the  writer  in  other  publications  (Jour.  Amcr.  Med.  Assoc, 
1002  (38),  220;  ibid.,  lOOO  (46),  341).  Ribbert  (Dent.  nicd.  WoH...  1<)03  (20). 
703)  has  also  advanced  a  similar  explanation  for  the  morplioloLncal  dilTcronces 
between  fatty  "de<j:('ncration"  and  ''infiltration,''  i.  e.,  that  the  dcixonerative 
changes  are  independcjit  of  fattv  accumulation. 

3oaJour.  Biol.  Chem.,  1017    (20),  310. 

30b  Fatty  cliangcs  in  the  viscera  niav  favor  tlicir  iriuisformalioii  into  adij)ocere 
(Miiller,  Vierteljalirs.  jrcM-icht.  Med.,  1015   (50),  251). 

37  Vierteljalirsch.  f.  gericht.  Med.,  1885  (42),  1. 


ADIPOCERE  411 

diflPusion  of  the  aimuoiiiiini  soaps  formed  during  the  decomposition.  As 
the  subcutaneous  fat  is  hardened  by  the  formation  of  soaps,  and  the 
bones  remain  to  hold  the  parts  in  position,  the  general  form  of  the 
body  is  preserved,  creating  the  impression  that  its  entire  su])stance  has 
been  converted  into  adipocere,  when  tlie  total  mass  may  actually  weigh 
but  twenty  pounds  or  so,  and,  according  to  Zillner's  estimate,  not  more 
than  one-tenth  of  the  muscle  substance  is  replaced  by  adipocere.  This 
false  impression  is  probably  responsible  for  much  of  the  mistaken 
idea  concerning  the  conversion  of  tissue  proteins  into  fatty  acids. 
Thus,  Schmidt  ^^  found  that  in  early  Egyptian  munniiies  60  per  cent, 
of  the  weight  of  the  lungs  and  30  per  cent,  of  the  spleen  consisted  of 
fatty  acids,  and  fell  into  the  usual  error  of  considering  this  conclu- 
sive evidence  of  transformation  of  proteins  into  fat. 

Numerous  attempts  have  been  made  to  prove  that  muscle  could  be 
thus  converted  into  fatty  acids  and  soaps,  but  although  success  has 
been  claimed  by  a  few,  the  results  are  not  entirely  convincing.^" 
Bacteria  can  convert  proteins  into  fats,  beyond  a  doubt,  and  they 
may  do  so  to  some  slight  extent  in  adipocere  formation,  but  probably 
this  factor  is  not  important. 

In  the  light  of  our  present  conception  of  fat  metabolism  it  is  prob- 
able that  the  process  of  adipocere  formation  occurs  as  follows:  The 
fatty  acids  of  the  fat  tissue  are  combined  by  the  ammonia  formed 
during  putrefaction,  removing  these  fatty  acids  from  the  normal  bal- 
ance of  fat  and  fatty  acids  in  the  fat  tissue;  as  a  result,  the  lipase  of 
the  fat  tissue  continues  to  split  the  fat,  and  more  fatty  acids  are 
produced,  which  likewise  go  to  form  soaps.  This  continues  until 
practically  all  the  neutral  fat  has  been  decomposed,  the  glycerol  dif- 
fusing rapidly  away.  The  soluble  soaps,  which  the  bacteria  do  not 
attack,  diffuse  into  the  softened  muscle  tissue,  w^hich  they  gradually 
replace  in  part.  In  the  meantime,  from  the  more  soluble  ammonium 
soaps,  calcium  and  magnesium  soaps  are  being  slowly  formed,  accord- 
ing to  the  usual  rule  of  double  decomposition  (that  the  least  soluble 
salt  will  be  formed  under  such  conditions)  ;  or  else,  if  an  acid  reaction 
develops,  free  fatty  acids  are  precipitated.  The  oleic  acid  seems  to 
be  converted  into  the  higher  fatty  acids  (Salkowski).*°  It  is  also 
possible  that  the  saponification  is  due  to  the  gradual  action  of  the  al- 
kaline fluids  produced  in  decomposition  of  the  tissues,  or  to  the  alka- 
linity of  the  water  in  which  the  body  lies.  Possibly  bacteria  may  be 
responsible  for  this  decomposition  of  the  fats  rather  than  the  body 
lipase,  for  Eijkman  *^  has  observed  that  certain  bacteria  growing  in 
fat-containing  agar  produce  calcium,  ammonium,  and  sodium  soaps, 
simulating  adipocere.^" 

s^Zeit.  all<;.  Physiol.,  1907   (7),  360. 

30  See  Rosenfeld.  Ercjeh.  der.  Physiol.,  Abt.  1.  1902  (1),  659. 
40  Festsdir.  f.  Virelmw,  1S91,  p.  23;  corroborated  bv  Schiitze. 
"Cent.  f.  Pakt..  1901   (29),  847. 

42  See  also  Covidalli.  Vierteliahrschr.  gerichtl.  :Med.,  1906  (32),  219;  and 
Schiitze,  Arch.  Hvg.,  -1912   (76),  116. 


412  RETh'OdJx'ESSJ  1  E  CHAyOES 

Zilliier "'  gives  the  folluwiiig  seheme  of  the  changes  that  take  place 
in  a  cadaver  undergoing  adipocere  formation:  (1)  Migration  of  fluid 
contents  of  the  body  (imbibition  of  blood  and  transudation) — one  to 
tour  weeks.  (2)  Decomposition  of  superficial  epidermis,  then  of 
corium — first  two  months.  (3)  Decomposition  of  muscle  and  gland 
parenchyma,  until  only  the  inorganic  part  of  the  bones  and  the  con- 
nective and  elastic  tissues  remain — three  to  twelve  months.  (4)  ]Mi- 
gration  of  neuti'al  fat,  ci-ystallization  and  partial  saponification  of  the 
higher  fatty  acids  in  the  panniculus ;  transformation  of  the  blood  pig- 
ment into  crystalline  form — four  to  twelve  or  more  months.*^ 

LIPEMIA 

Normally  the  blood  contains  a  considerable  amount  of  fats  and 
lipoids,  varying  somewhat,  but  not  greatly,  with  the  diet.  The  older 
literature  gave  figures  varying  widely,  but  analyses  by  more  modern 
methods  ^*  give  figures  for  the  ether-soluble  constituents  of  the  normal 
plasma  (before  breakfast)  ranging  ordinarily  from  0.57  to  0.82  per 
cent.,  of  which  cholesterol  and  phosphatid  are  about  equal  (0.2  to  0.3 
per  cent.)  with  very  little  neutral  fat  (0.1  to  0.2  per  cent.).  In 
various  diseases,  exclusive  of  diabetes,  the  total  lipin  content  was 
found  by  Bloor  to  be  about  normal,  but  the  proportion  of  the  dif- 
ferent lipins  varied  somewhat.  After  taking  fat-rich  food,  however, 
there  may  be  a  considerable  excess  of  the  food  fats  in  the  serum,  and  it 
is,  therefore,  extremel.y  difficult  to  say  just  when  the  amount  of  fat  in 
the  blood  is  large  enough  to  be  considered  as  a  lipemia,  especially 
since  after  every  fatty  meal  there  is  enough  fat  in  the  blood  to  make 
it  turbid.'*^  B.  Fisher  ■**  states  that  we  may  speak  of  a  pathological 
lipemia  when  we  have  a  distinctly  cloudy  blood  or  serum,  which  is 
clarified  by  shaking  with  ether  through  the  dissolving  out  of  fat  which 
can  then  be  separated  from  the  ether.  Earlier  writers  described,  in- 
correctly, lipemia  in  many  conditions,  but  recent  writers  mention  it 
chiefly  as  occurring  in  alcoholism  and  diabetes.  By  far  the  greatest 
amounts  of  fat  are  observed  in  the  latter  condition,  and  diabetic 
lipemia  is  always  accompanied  by  an  acidosis,  although  acidosis  often 
occurs  without  lipemia.  Experimental  pancreatic  diabetes  may  be 
accompanied  by  lipemia.*"  Neisser  and  Dei-lin''"  found  19.7  per  cent, 
of  fat  in  the  blood  of  a  patient  with  dial)etie  coma  (after  death  24.4 
per  cent,  was  found)  whose  urine  contained  0.8  per  cent,  of  fat,  and 
through  analysis  of  this  and  other  material  came  to  the  conclusion  that 
the  fat  comes  directly  from  the  chyle;  i.  c,  tliat  it  is  food  fat,  not 

*3  Sclerema  neonatorum  is  caused  by  hardening  of  tlic  subcutaneous  fat.  be- 
cause of  a  low  proportion  of  oleic  acid.  ( Bcvcr.  ^'crll.  Dcut.  Path.  Gcscll.,  IHOS 
(12),  305.) 

4-«  Bloor,  Jour.  IJiol.  Cliciii.,   litlfl    (iZf)).  577. 

4"  Neisser  and  Braiuiin^r,  Zeit.  oxp.  Patli.  u.  Tlier..  1907    (4).  747. 

"♦s  Virchow's  Arc'li..  lOO."}   (172),  ."^O.     Kcsunie  and  ('(unplctc  literature. 

"»!».Seo,  Ardi.  cxp.  Path.  u.  Pliariii.,  liKMi    (dl  ),   1. 

J^oZeit.  klin.  Med.,  1904    (51),  428. 


ij/'inuA  413 

l)ody  fat.  Fisclier  found  an  average  of  18.129  per  cent,  in  his  case, 
including  at  least  0.478  per  cent,  of  cholesterol,  with  no  lipuria  and 
very  small  amounts  of  fatty  acids;  of  the  fat,  about  67.5  per  cent,  was 
olein.  As  higii  as  27  per  cent,  of  fat  in  the  blood  has  been  found."'-  In 
many  cases  the  increase  is  chiefly  in  tlie  lipoids,  lipoidemia,^^  and  in 
acidosis  there  is  said  to  be  an  especial  increase  in  cholesterol  (Adler).^* 

It  is  an  important  question  whetlier,  witli  such  high  (luantities  of  fat 
in  the  blood,  fat  embolism  ma}'  result,  for  it  is  possible  that  at  least 
some  of  the  cases  of  diabetic  coma  are  due  to  such  fat  embolism  in 
the  cerebral  vessels.  Ebstein  ''^  considers  this  a  possible,  but  not  a 
common,  occurrence,  because  the  droplets  are  too  small  to  cause  oc- 
clusion of  the  vessels  unless  they  combine  to  form  large  droplets. 
Fischer  doubts  if  the  droplets  ever  fuse  together  enough  to  cause  em- 
bolism, supporting  his  contention  both  by  experiments  and  clinical 
records,  but  cases  have  been  reported  as  fat  embolism  from  diabetic 
lipemia.''^ 

The  cause  of  lipemia  has  not  yet  been  satisfactorily  determined.  In 
alcoholism  it  is  commonly  ascribed  to  a  failure  to  burn  fat,  because 
of  the  presence  of  the  more  readily  oxidized  alcohol,  and  the  common 
coexistence  of  diabetes  and  lipemia  suggests  for  both  a  common  cause ; 
i.  e.,  lack  of  oxidation  of  fat  and  sugar.  In  corroboration  may  be 
cited  the  occurrence  of  lipemia  in  other  conditions  associated  with 
defective  oxidation ;  /.  e.,  pneumonia,  anemia,^**  phosphorus-poisoning. 
As  we  are  still  unfamiliar  with  the  essential  factors  and  steps  in  the 
oxidation  of  fat,  it  would  be  mere  speculation  to  attempt  to  explain 
further  the  reason  for  the  failure  of  destruction  of  the  fat.  The 
origin  of  the  fat  in  lipemia  is  likewise  undetermined.  Ebstein  con- 
siders that  it  arises  partly  from  the  food,  partly  from  fattj'  degenera- 
tion of  the  cells  of  the  blood,  the  vessel-walls,  and  the  viscera.  Neisser 
and  Berlin  consider  it  as  merely  food  fat  coming  from  the  chyle  and 
accumulated  in  the  blood.  Fischer  believes  that  it  is  largely  derived 
from  the  fat  depots,  and  that  because  of  loss  of  the  lipolytic  power 
of  the  blood  it  cannot  be  rendered  diffusible,  and  hence  it  cannot  enter 
the  tissues  where  it  is  normally  consumed.  Sakai  ^"'^  also  found  a  low 
lipase  content  in  the  blood  and  suggests  that  fat  entering  the  blood  is 
unable  to  leave  it  because  of  defective  lipolysis.  Klemperer  and  Um- 
ber hold  that  it  comes  from  disintegration  of  tissue  cells,  but  are 
unable  to  determine  the  cells  concerned. 

Bloor's  studies^""  support  strongly  the  view  that   the   fats  come 

52  Frugoni  and  ^ilarchetti,  Berl.  klin.  Woch.,  1908   (4.")),  1S44. 

53  See  Weil,  IMiineh.  med.  Woch.,  1012   (59),  2096. 
■'•••iBerl.  klin.  Woch.,  1910    (47),  132.3. 

53  Virchow's  Arch.,  1899  (1.55),  571. 
51  Hedren,  Svenska  Liik.  Handl.,  1910    (42),  933. 

56  See  Boggs  and  Morris    (Jour.  Exper.  Med.,    1909    (11),  553),   wiio  produced 
lipemia  bv  repeatedly  l)leeding  rabbits. 
ssaBioo'hem.  Zeit.,'l914    (62).  387. 
56b  Jour.  Biol.  Chem.,  1916    (26),  417. 


414  RETROGRESSIVE  CHANGES 

from  the  food,  for  he  found  lipeuiia  oul}'  in  diabetics  receiving  fat  in 
their  food,  and  under  fasting  an  existing  lipemia  disappears.  Choles- 
terol increases  parallel  Avith  the  fat,  while  lecithin  is  relatively  little 
increased.  In  severe  diabetes  without  lipemia  the  lipins  are  ail  much 
increased  in  the  plasma,  but  with  the  relative  proportions  about  as  in 
normal  individuals,  although  with  a  tendency  for  the  fats  to  accumu- 
late in  excess.  The  facts  that  fat  oxidation  depends  upon  carbohy- 
drate oxidation,  and  also  that  in  diabetics  excessive  fat  feeding  is 
usual,  are  probably  significant  in  the  causation  of  diabetic  lipemia. 

PATHOLOGICAL  OCCURRENCE  OF  FATTY  ACIDS 

Fatty  acids  occasionally  occur  free  in  pathological  processes.  The 
l)est  example  of  this  is  fat  necrosis  {q.  v.),  where  cr3-stals  of  fatty 
acids  appear  in  the  necrotic  fat-cells,  arising  through  splitting  of  fat, 
and  later  becoming  combined  with  calcium  from  the  blood.  Similar 
crystals,  consisting  of  a  mixture  of  palmitic  and  stearic  acids,  fre- 
iiuently  called  margarin  or  margaric  acid  crystals,  may  be  found  in 
decomposed  pus,  in  sputum  from  bronchiectatic  cavities  and  from 
gangrene  of  the  lungs,  in  gangrenous  tissue,  and  in  atheromatous 
areas.  According  to  Schwartz  and  Kayser,^^  the  free  fatty  acids,  at 
least  in  pulmonary  gangrene,  arise  from  lipolysis  by  bacterial  action 
rather  than  by  the  lipase  of  the  tissues.  Eichhorst  found  crystals  of 
fatty  acids  in  the  neighborhood  of  acute  patches  of  sclerosis  in  the 
central  nervous  system  in  multiple  sclerosis,  and  McCarthy  ^^  found 
them  in  a  spinal  cord  undergoing  secondary  degeneration  from  com- 
pression. Whipple  ^°  describes  a  case  with  deposits  of  fatty  acids  and 
neutral  fat  in  the  w^all  of  the  intestine  and  the  mesenteric  glands, 
while  soaps  and  fatty  acids  are  said  to  be  present  in  excess  in  chronic 
appendicitis."**  Soaps  and  fatty  acids,  especially  oleic  acid  and  oleates, 
are  highly  toxic,  and  their  profound  hemolytic  power  has  been 
thought  of  importance  in  pathological  conditions,  especially  bothrio- 
cephalus  anemia."^  (See  Hemolysins,  Chap,  viii.)  The  fatal  dose  of 
sodium  oleate  for  rabbits  is  0.15  gm.  per  kilo  (Leathes),  The  salts 
of  higher  fatty  acids  above  capric  are  hemolytic,  while  those  from 
caproic  down  are  not,  nonoic  acid  salts  being  the  turning  point 
(Shimazono).''-  The  toxicity  of  soaps  may  be  related  to  their  marked 
power  to  inhibit  proteolytic  enzymes.'^-'^ 

The  fatty  acids  may  be  stained  green  by  copper  acetate,  according 
to  Benda's  method,  and  if  then  treated  with  hematoxylin,  they  turn 
black."^     With   Nile   blue   sulphate  they   stain  blue,   forming   a  blue 

57Zeit.  klin.  ^Mcd.,  1905   (56),  111. 

58  Univ.  of  PtTin.  Med.  15ull.,  190:?    (10),  141. 

50  Bull,  .lohiis  Hopkins  Ilosp.,  1!I07   (18),  l^S'l. 

8'>  .Anthony,  .lour.  Mod.  Itos.,  1911    (20),  .S5!). 

ci  Faust,  Suppl.  V,d..  ScliniicdclxTfi's  Arch.,  lOOS,  p.   171. 

•••2  Z.  Inimunitiit.,  Kef.,  1!)11    (4),  (i.")6. 

"sa.Joblinp  and  Petersen,  Jour.  K.\p.  Mod.,  1014   (10),  251. 

03  Fischler,  Cent.  f.  Path.,  1004   (15),  013. 


I'ATIlOLOalVAL  OVCUh'KEXCE  OF  CHOLKSTEROL  415 

salt,  while  the  neutral  fats  are  stained  red  by  the  oxazone  base  (J.  L. 
Smith).  Fischler  and  Gross"''  state  that  fatty  acids  are  present  in 
atheromatous  areas  and  about  the  margin  of  anemic  infarcts,  but 
are  not  recognizable  by  this  method  in  such  fatty  degenerations  as 
])neuin()nic  exudates,  caseation,  etc.  Klotz  "^  considers  that  calcium 
soaps  are  formed  as  the  first  step  in  pathological  calcitication,  accord- 
ing to  microchemical  evidence ;  but  a  chemical  investigation  of  the 
same  question  did  not  give  tlie  writer  positive  results.^'  In  fatty 
cells,  especially  in  the  liver,  crystals  are  often  found  and  interpreted 
as  fatt\-  ac'ds,  whirli  hre  really  crystals  of  neutral  fats."' 

PATHOLOGICAL  OCCURRENCE  OF  CHOLESTEROL 's 

Cholesterol  in  crystals  is  found  under  somewhat  the  same  conditions 
as  the  fatty  acids,  and  although  cholesterol  is  not  a  fat,  but  an  alco- 
hol, its  phj'sical  properties  are  so  similar  that  it  may  be  considered 
in  this  place.  (See  "Gall-stones,"  Chap,  xv,  for  further  discvission.) 
The  characteristic  large  flat  plates  of  cholesterol  may  be  found  in  any 
tissue  in  which  cells  are  undergoing  slow  destniction,  and  where  absorp- 
tion is  poor.  Therefore,  they  are  found  frequently  in  atheromatous 
patches  in  the  blood-vessels,  encapsulated  caseous  areas,  old  infarcts 
and  hematomas,  inspissated  pus-collections,  dermoid  cysts,  hydrocele 
fluids,  etc. ;  especially  large  amounts  occur  in  the  cholesteatomatous 
tumors  of  the  ear  and  cranial  cavity. *'*' 

In  degenerative  conditions  of  the  central  nervous  system  ^^^  choles- 
terol may  be  present  in  the  spinal  fluid  (Pighini'°),  and  in  an  old 
pleural  effusion  as  much  as  1.25  per  cent,  of  cholesterol  has  been  found 
(Ruppert  "^).  Windaus  ''-  found  that  normal  aortas  contain  about  0.15 
per  cent,  cholesterol,  while  in  two  atheromatous  aortas  he  found  1.8 
per  cent,  and  1.4  per  cent.,  the  increase  being  more  in  the  cholesterol 
esters  than  in  the  free  cholesterol.  Amyloid  kidne,ys,  however,  show  an 
increase  only  in  the  cholesterol  esters,  and  not  at  all  in  the  free  choles- 
terol. (See  Relation  of  Lipoids  to  Fatty  Metamorphosis,  p.  404.) 
Ameseder  ^^  found  that  28.56  per  cent,  of  the  ether  extract  of  athero- 
matous aortas  was  cholesterol.  The  claim  of  Chauffard  that  arcus 
senilis,  xanthelasma,  and  other  ocular  conditions  depend  on  choles- 
terol deposition  is  not  substantiated  by  Mawas."*     In  liquids  the  crys- 

GiZiegler's  Beitr.,  lOO.l   (7th  suppl.),  343. 
G5  Jour.  Exp.  :\rocl.,  100.5    (7),  033. 

66  Wells,  Jour.  Med.  Research,  190G  (14).  491. 

67  Smith  and  White,  Jour.  Path,  and  Bact.,  1907    (12),  126. 

68  Concerning  the  chemistry  of  cholesterol  see  introductory  chapter. 

69  See  Bostrocm,  Cent.  f.  Path.,  1897   (8),  1. 

69a  Southard  has  described  cholesterol  concretions  up  to  2  cm.  diameter  in  the 
brain  and  cord.      (Joiir.  Amer.  Med.  Assoc,  1905   (45),  1731.) 
ToRiforma  Med..   1909    (25),  67. 
7iMiinch.  med.  Woch.,  1908   (55),  510. 
T2Zeit.  physiol.  Chem.,  1910    (67),   174. 
73Zeit.  physiol.  Chem.,  1911    (70),  458. 
T4Monatsbl.  f.  Augenheilk.,  1912    (13),  604. 


416  RETROGRESSIVE  CHANGES 

tals  form  <i'listeiiing  scales ;  in  fresh  tissues  tliey  may  be  recognized  by 
their  solubility  in  ether,  chloroform,  hot  alcohol,  etc.,  and  by  their 
color  reactions.  In  histological  specimens  prepared  by  the  usual 
methods  the  cholesterol  is  dissolved  out,  but  the  resulting  clear-cut 
clefts  are  quite  characteristic.  In  fresh  specimens  in  which  choles- 
terol crj'stals  are  present,  on  treatment  with  five  parts  concentrated 
sul])huric  acid  and  one  of  water,  the  edges  of  the  crystals  become 
carmine  red,  then  violet.  Concentrated  sulphuric  acid  plus  a  trace 
of  iodin  colors  the  crystals  in  sequence,  violet,  blue,  green,  and  red. 
Hirschsohn  '^^  recommends  a  reaction  with  a  90  per  cent,  solution  of 
trichloracetic  acid  in  HCl,  which  gives  red,  then  violet,  then  blue.  The 
results  of  microehemieal  examination  are  said  not  to  agree  at  all  quan- 
titatively Avith  analytic  results."^''' 

Since  all  cells  contain  cholesterol,''^  it  is  perhaps  accumulated  as  one 
of  the  least  soluble  products  of  their  disintegration.  The  origin  of 
the  normal  cell  cholesterol  is  unknown,  but  that  which  is  liberated  by 
normal  disintegration  of  cells  seems  to  be  retained  and  worked  over." 
It  is  not  destroyed  during  autolysis."  Cholesterol  is  generally  con- 
sidered, but  without  convincing  proof,  to  be  a  product  of  protein  de- 
composition ;  if  this  is  true,  then  the  cholesterol  found  in  disintegrat- 
ing tissues  may  be  formed  from  the  cell  proteins  during  their  de- 
composition.'^'* Apparently  cholesterol  crystals  may  be  slowly  re- 
moved, the  chief  factor  probably  being  the  giant-cells  that  are  often 
found  surrounding  them,®°  and  the  large  "foamy"  endothelial  cells 
that  take  up  especially  the  uncrystallized  cholesterol.  In  general  they 
behave  as  inert  foreign  bodies.  Xanthomatous  masses  of  various  kinds 
all  seem  to  be  composed  of  deposits  of  cholesterol  esters  which  lead  to 
proliferative  and  phagocytic  reactions  in  the  fixed  tissues.^*"^ 

Cholesterolemia.**"" — Normal  blood  contains  1.7  to  2.5  per  mille  of 
cholesterol,  of  which  about  55  per  cent,  is  in  the  corpuscles,  both  in 
normal  and  pathological  conditions  (Bacmeister  and  Henes**"^). 
Cholesterol -rich  diet  causes  a  slight  increase,  but  a  more  marked  in- 
crease is  said  to  be  obtained  in  pregnancy,  nephritis,  early  arterio- 
sclerosis, obesity,  diabetes,  and  obstructive  jaundice.     According  to 

'SPharm.  Contrallialle,   in02    (4.3).  357. 

v-.a  Thavsen,  Cent,  allfj.  PatlioL,  101.')    (2fi),  433. 

TO  See  boree.  Bioeliom.  Jour.,  1009    (4),  72. 

"Ellis  and  Cardiier,  I'roc.  Royal  Soc,  London.   1012    (S4).  401. 

Ts  Corpcr,  Jour.  Hiol.  Cliem., '  1012  (11),  37:  Shiliata,  nioclioni.  Zoit.,  1011 
(31),  321. 

70  Of  historical  interest  is  Austin  Flint's  idea  that  clioiesterol  in  tlie  hlood  is 
an  important  factor  in  into.xications,  especially  in  icterus  (Anier.  Jour.  Med. 
Sci.,  1862  (44),  20).  All  recent  evidence  is  tothe  etTect  that  cholesterol  is  not 
toxic. 

so  See  LeCount,  .Tour.  iNled.  Research,  1002  (7),  liKl;  ('or|)cr,  .lour.  K\p.  ^fcd., 
101.-)    (21).  170:   Stewart.  .Toin-.  Patli.  and  Ract..   lOlfj    (10),  30.'). 

><""  Lit.'rature  -riven  1)V  Hosenl)looni,   Arcli.  Int.  :\Ied.,   1013    (12).  30,-). 

><'>b  Pil,li,,trnipliv  hv  licwcv.  Arch.  Tut.   .Med.,   101(5    (17),  7r>7. 

8o<- l)<'iit     iiicil.  'W.irh..    l!il';!    ( :!".»),  r>44. 


AMYLOID  417 

some  observations,  in  nepliritis  the  amount  of  cholesterol  beare  no  re- 
lation to  tlie  albuminuria,  and  in  uremia  it  may  be  low;  acute  febrile 
diseases  usually  show  a  lowered  cholesterol,  which  is  unchanfjed  in 
tuberculosis.  The  blood  content  has  been  reported  as  low  in  febrile 
cutaneous  diseases,  but  high  in  afebrile  cutaneous  diseases  associated 
with  eosinoi)hilia/"''  However,  Denis  -'-"^  states,  after  examination  of 
a  large  number  of  cases,  that  hypercholesterolemia  was  found  only  in 
diabetes,  and  that  low  cholesterol  values  are  found  in  cachexia  or  pros- 
tration, but  are  not  characteristic  of  any  particular  disease. 

Experimental  hypercholesterolemia  in  animals  leads  to  a  deposition 
of  cholesterol  in  various  organs,  especially  the  aorta,  kidneys  and 
liver,  accompanied  by  degeneration  in  the  parenchymatous  structures, 
and  excretion  of  cholesterol  in  the  urine  and  bile ;  gall  stones  may  be 
formed  (Dewey).  Sometimes  lipoid-filled  endothelial  cells  become  so 
abundant  in  the  spleen  as  to  resemble  Gaucher 's  disease  (Anichkov, 
]\Ic^Ieans  ~"").  Cholesterol  in  the  blood  reduces  phagocytic  activity 
and  antibody  formation  in  experimental  animals.^°^ 

The  ratio  of  free  cholesterol  to  cholesterol  esters  in  normal  human 
blood  is  nearly  constant,  the  esters  being  about  33.5  per  cent,  in  the 
blood  and  58  per  cent,  in  the  plasma ;  in  pregnancy  the  proportion  of 
cholesterol  esters  is  high,  in  cancer  and  nephritis  it  is  low.®°^ 

AMYLOID  «i 

Virchow,  in  1853,  made  the  first  study  of  the  nature  of  the  substance 
characteristic  of  "lardaceous"  degeneration,  and  considered  it  to  be 
a  sort  of  animal  cellulose,  because  it  often  became  blue  if  treated  with 
iodin  followed  by  sulphuric  acid.  To  this  resemblance  in  staining 
reaction  we  owe  the  unfortunate,  misleading,  but  generally  used,  name 
amyloid.^-  It  was  but  a  few  years  (1859)  before  Friedreich  and 
Kekule  showed  that  the  substance  in  question  was  of  protein  nature ; 
their  methods  were  very  crude,  but  the  main  fact  was  soon  better 
substantiated  by  KiOine  and  Rudneff  (1865).     Krawkow,*^  however, 

sodFischl.  Wien.  klin.  Woch.,  1914   (27),  982. 

soeJour.  Biol.  Chem.,  1917    (29),  9.3. 

80f  Jour.  Med.  Res..  1916    (.33),  481. 

80g  Dewey  and  Nnzuni,  Jour.  Infect.  Dis.,  1914    (In).  472. 

80h  Bloor  and  Knudson,  Jour.  Biol.  Chem.,  1917    (29).  7. 

81  General  literature  to  1893,  see  Wichmann,  Ziepler's  Beitr..  1893  (13).  487: 
also  Lubarsch,  Ergeb.  allg.  Path.,  1897  (4),  449;  discussion  in  the  Verb.  Deut. 
Path.  Gesellsch..  1904  (7),  2-,51:  Davidsohn.  Virchow's  Arch.,  1908  (192),  226, 
and  Ergebnisse  allg.  Path..  1908    (12),  424. 

82  In  view  of  the  fact  that  this  substance  is  cheniically  related  to  chondrin, 
and  that  it  also  closely  resembles  this  substance  physically,  it  has  seemed  to  the 
writer  that  the  name  "chondroid"  would  be  inuch  more  appropriate  than  any  of 
the  many  more  or  less  misleading  and  inappropriate  titles  tliat  are  at  present  in 
use.  The  very  multiplicity  of  these  terms,  however,  prohibits  any  attempt  to 
introduce  still  another.  A  particularly  unfortunate  source  of  confusion  exists 
in  the  use  of  the  name  amyloid  for  a  vegetable  substance,  formed  by  the  action 
of  acids  upon  cellulose. 

83  Arch.  exp.  Path.  u.  Pharm.,  1897   (40),  196. 

27 


418  RETROGRESSIVE  CHANGES 

in  1897  g-ave  us  the  first  ^ood  idea  of  the  composition  of  amyloid  sub- 
stance through  his  amplification  of  Oddi's®^  observation  that  amyloid 
organs  contain  chondroitin-sulphuric  acid,  finding  that  amyloid  is  a 
compound  of  protein  with  this  acid,  similar  to  nucleoprotein,  which 
is  a  compound  of  nucleic  acid  and  protein.  This  work  has  received 
general  acceptance,  although  a  later  paper  by  Hanssen  ®^  reports  a 
study  of  amyloid  material  isolated  in  pure  condition  from  sago  spleens 
by  mechanical  means,  which  contained  no  chondroitin-sulphuric  acid, 
although  the  amyloid  organs  taken  i)i  toto  do  contain  an  excess  of  sul- 
phur as  sulphate.  This  important  contradiction  to  prevailing  ideas 
has  not,  so  far  as  I  can  find,  been  subjected  to  investigation  by  others, 
with  the  exception  of  a  casual  remark  by  Mayeda  ^^  that  a  prepara- 
tion of  amyloid  which  he  had  made  did  not  yield  sulphuric  acid. 

Chondroitin-sulphuric  acid,  which  has  been  studied  especially  hy  jMorner  and 
hy  Sclimicdeborg,8"  has  tlie  formula  C\sH^^NS0,7,  accordin<T  to  tlie  latter,  and 
yields  on  cleavage  chondroitin  and  sulphuric  acid,  as  follows: 

C,sHj,NSOiT  +  H,0  =  CigHjjNO,,  +  USO, 

Kondo,88  however,  gives  it  an  empirical  formula  of  Ci^HojNSOia,  there  being  ap- 
parently two  equivalents  of  tlie  base  for  each  SO4  group.  Levene  and  La  Forge  «' 
have  demonstrated  that  cliondroitin-sulphuric  acid  consists  of  sulphuric  acid, 
acetic  acid,  cliondrosamine  wliich  is  an  isomer  of  glucosamine,  and  glucuronic  acid. 
It  unites  with  histones  and  forms  a  precipitates^  Chondroitin  is  a  gummy  sub- 
stance which  in  turn  may  be  split  into  acetic  acid  and  a  reducing  substance, 
chondrosin.  Chondroitin-sulphuric  acid  is  the  characteristic  component  of  car- 
tilage, but  it  is  also  found  in  mucin  (Levene),  and  in  the  walls  of  the  aorta  and 
other  elastic  structures  (Krawkow).  It  has  also  been  found  in  a  uterine  fibroma 
and  in  bone  tissue  by  Krawkow,  but  could  not  be  found  in  the  parenchymatous 
organs,  normal  and  pathological,  or  in  chitinous  structures.  Morner  has  also 
found  it  in  a  chondroma. 

Chemistry  of  Amyloid. — Krawkow  separated  amyloid  from  nu- 
cl('oi)rotein,  to  which  it  is  most  closely  related,  by  dissolving  both  sub- 
stances from  the  minced  amyloid  organs  with  ammonia,  precipitating 
with  acid,  and  then  taking  up  the  amyloid  with  Ba(0H)2  solution,  in 
which  the  nucleoprotein  does  not  dissolve.  Amyloid  thus  isolated  is 
a  nearly  white  powder,  which  is  easily  soluble  in  alkalies,  but  slightly 
in  acids,  and  is  very  resistant  to  pepsin  digestion.  The  elementary 
composition  was  found  by  Krawkow  to  be  approximately  as  follows : 

C  =  40-50%  ;  H  =  6.65-7%  ;  N  =  13.8-14%  ;  S  =  2.65-2.!)%  ;  P  in  traces  only. 

Quite  similar  analytic  results  have  been  obtained  by  Neuberg,"*^ 
who  coi-i'oboi'ated  Krawkow 's  finding  of  a  body  of  apparently  similar 

Si  Ibid.,  1894   (.'?.3),  .377. 

sf.  Biochem.  Zeit.,  1008    (13),  1S5. 

80  Zeit.  phvsiol.  Chem.,  1000    (58),  475. 

87  Morner."  Skand.  Arch.  Divsiol.,  1880  (1).  210;  Zeit.  phvsiol.  Ciiem.,  1805 
(20),  357,  and  1807  (23),  311;  Schniiedeberg,  Arcli.  exp.  rath.  u.  rharm..  1801 
(28),  358.  See  also  Levene  and  La  Forge,  Jour.  Biol.  Chem.,  1013  (15),  60  and 
155;   1014   (18),  123. 

88  Biochem.  Zeit.,  1010   (26),  116. 

89  Pons,  Arch,  internat.  phvsiol.,  1000    (8),  303. 
00  Verb.  Deut.  Path.  Gesell.",  1004   (7),  19. 


AMYJJJJD 


419 


composition  in  the  normal  aorta.  Neuberg  has  studied  especially  tlie 
protein  constituent  of  the  amyloid  compound,  and  found  it  character- 
ized by  a  high  proportion  of  diamino-nitrogen,"^  as  compared  with 
most  proteins,  as  shown  in  the  following  table  giving  the  percentage 
of  the  total  X  contained  in  each  of  the  three  forms,  amid-nitrogen 
(ammonia),  monamino-acids,  and  diamino-acids: 

Table  I 


Liver  amyloid 
Spleen  amyloid 
Aorta   "amyloid" 
Gelatin 
Casein 


Monamino- 

Diamino- 

acid 

acid 

Amid 

nitrogen 

nitrogen 

nitrogen. 

43.2 

51.2 

4.9 

30.6 

57.0 

11.2 

54.9 

36.0 

8.8 

62.5 

35.8 

1.6 

76.0 

11.1 

13.4 

The  variations  in  the  composition  of  the  different  amyloids,  as  shown 
in  the  above  table,  indicate  that  the  protein  group  may  vary  in  dif- 
ferent organs  in  different  cases,  and  also  indicate  that  the  "amyloid- 
like" substance  of  normal  vessels  is  not  the  same  as  the  pathological 
substance.  Corresponding  variations  were  found  in  the  apportion- 
ment of  the  sulphur  between  that  which  is  in  the  form  of  oxidized  sul- 
phur and  the  unoxidized  sulphur.  The  proportion  of  the  different 
amino-aeids  in  the  protein  constituent  of  amyloid  is  strikingly  like  that 
of  thymus  histon,  and  entirely  dissimilar  to  the  apparently  closely 
related  elastin,  as  shown  by  Table  II, 


Table  II 

Cleavage  products  (in  percentages) 

Amyloid 

Elastin 

Thymus 
histon 

filvcocoll 

0.8 

25.8 

0.5 

Leucine     

22.2 

45.0 

11.8 

(ilutaminic   acid 

3.8 

0.7 

3.7 

Tyrosine         

4.0 

0.3 

5.2 

a-Proline 

3.1 

1.7 

1.5 

Arginine 

13.9 

0.3 

14.5 

Lysine .'      . 

11.6 

7.7 

This  carries  out  the  resemblance  of  amyloid  to  nucleoproteins,  and, 
likewise,  Neuberg  found  amyloid  very  slowly  digested  by  pepsin,  and 
much  better  by  trv-psin,  although  less  rapidly  than  simple  protein;  it 
is  also  destroyed  by  autolytic  enzymes,  for  amyloid  tissues  readily 

91  Corroborated  by  Jackson  and  Pearce  (Jotir.  Exp.  Med.,  1907  (9).  520),  but 
not  by  Mayeda  ( Zeit.  physiol.  Chem.,  1909  (58),  469),  who  found  histidine,  which 
Neuberg  had  missed,  and  a  lower  arginine  and  lysine  content  than  histon  re- 
quires. 


420  RETROGRESSIVE  CHANGES 

undergo  autolysis."-  Neuberg  considers,  from  the  above  results,  that 
amyloid  is  probably  a  transformation-product  of  the  tissue  protein, 
similar  to  the  transformation  of  simple  proteins  into  protamins  that 
occurs  in  the  testicle  of  spawning-  salmon  as  they  go  up  the  streams, 
as  shown  by  Micscher's  classical  studies.  Raubitschek  "^  found  that 
isolated  amyloid,  when  used  for  immune  reactions,  behaved  like  a 
specific  protein,  different  from  the  normal  proteins  of  the  animal  from 
whence  it  came  and  apparently  biologically  the  same  in  different  spe- 
cies.     (This  observation  awaits  confirmation.) 

Krawkow  considers  that  amyloid  differs  from  normal  chondroitin- 
sulphuric  acid  compounds,  such  as  cartilage,  in  that  in  the  latter  the 
acid  radical  is  in  a  loose  combination  with  the  protein,  while  in  amy- 
loid the  combination  is  a  very  firm  one,  perhaps  in  the  nature  of  an 
ester.  The  occurrence  of  the  typical  amyloid  reaction  in  what  ap- 
pears otherwise  to  be  normal  cartilage,  occasionally  observed  in  senile 
tissues,  may  be  due  to  the  transformation  of  loosely  bound  into  firmly 
bound  chondroitin-sulphuric  acid.  In  any  event,  amyloid  is  not  essen- 
tially a  pathological  product,  but  rather  a  slightly  modified  nonnal 
constituent  of  the  body.  However,  in  view  of  the  contradictory  results 
of  Hanssen  and  Mayeda,  as  yet  uncontroverted,  the  chemical  nature  of 
amyloid  must  be  considered  as  undetennined.  An  important  con- 
sideration is  that  amyloid  deposition  occurs  under  similar  conditions 
in  all  sorts  of  animals,  including  birds ;  it  is  very  often  found  in 
the  livers  of  antitoxin  horses,  and  mice  are  especially  prone  to  a 
severe  amyloidosis  after  relatively  slight  and  brief  infectious  pro- 
cesses."* 

Staining  Properties. — The  classical  reaction  for  amyloid  is  its 
staining  a  reddish  brown  when  treated  with  iodin  (best  as  Lugol's  so- 
lution) in  the  fresh  state.  Such  stained  specimens,  if  afterward 
treated  with  dilute  sulphuric  acid,  usually  become  blue  or  greenish,  but 
may  merely  tuni  a  deeper  brown.  Occasionally  old  compact  amyloid 
may  stain  bluish  or  green  with  iodin  alone.  The  iodin  reaction  dis- 
appears in  specimens  that  have  been  kept  for  some  time  in  preserv- 
ing fluids,  or  in  tissues  that  have  become  alkaline,  and  is  generally 
less  persistent  than  the  metachromatic  staining  by  methyl-violet  or 
methyl-green,  which  color  the  amyloid  red.  Occasionally  an  otherwise 
typical  amyloid  will  fail  to  react  to  iodin,  but  will  stain  well  with 
methyl-violet.  All  these  variations  may  occui-  in  different  specimens 
from  the  same  body,  and  the  blue  iodin-sul]ihuric  acid  reaction  is 
usually  given  well  only  by  splenic  amyloid.  These  variations  proba- 
bly depend  upon  the  age  and  stage  of  development  of  the  amyloid,  or 
upon   secondaiy   alterations,   and    are  perhaps   related    to   Neuberg's 

''2  f 'oncorniTiL,'  llic  ubsorption  of  aiinloid  soe  Dantcliokdw,  Virclunv's  Arcliiv., 
1907    nS7),  1. 

osVorh.  Dont.  Path.  Cosoll..  1010   (14),  27.1 

niSoo  Fin/.i,  I^  Rporinifiit..  1011  {C>r^).  4S.1 ;  Davidsohn,  VirchowV  Arcli..  1008 
(192),  226. 


THE  ORIGIN  OF  AMYLOID  421 

observations  on  the  difference  in  composition  of  amyloid  of  different 
origins. 

Krawkow  studied  these  reactions  witli  pure,  isolated  amyloid,  and 
found  evidence  that  the  iodin  reaction  depends  upon  the  physical 
properties  of  the  amyloid,  while  tlie  methyl-violet  stain  is  a  chemical 
reaction,  and  hence  the  iodin  reaction  is  much  the  more  readily  altered 
or  lost.  As  Dickinson  ''^  says,  amyloid  stains  with  iodin  simply  as 
if  it  absorbed  the  iodin  more  than  does  the  surrounding  tissue.  The 
methyl-violet  reaction  is  due  to  the  dye  forming  a  compound  with  the 
ehondn)itin-sali)huric  acid,  for  Krawkow  found  that  these  substances 
unite  witli  one  another  to  form  a  rose-red  precipitate.  Hanssen,  how- 
ever, holds  tliat  the  dyes  react  with  the  protein,  the  iodin  with  some 
other,  unknown  labile  substance.  Schmidt  found  that  implanted 
pieces  of  amyloid  lost  their  iodin  reaction  as  they  underwent  auto- 
lysis, while  the  methyl-violet  reaction  was  still  very  distinct.'*'^  It 
is  evident,  therefore,  that  iodin  is  not  by  itself  a  specific  stain  for 
amyloid,  especially  as  glycogen  gives  a  similar  reaction,''"  while  true 
amyloid  may  not  react. 

THE   ORIGIN  OF  AMYLOID 

This  question  has  not  been  at  all  cleared  up  as  j^et  by  the  advances 
made  in  our  knowledge  of  the  chemistry  of  amyloid  substance.  The 
fact  that  chondroitin-sulphuric  acid  is  a  characteristic  constituent 
suggests  that  this  body  may  be  liberated  in  considerable  amount  dur- 
ing the  destructive  processes  to  which  amyloidosis  is  usually  sec- 
ondary: this  idea  is  further  supported  by  the  fact  that  amyloidosis 
occurs  particularly  after  chronic  suppuration  in  bone  and  lungs,  both 
of  which  tissues,  according  to  Krawkow,  contain  chondroitin-sulphuric 
acid.  This  idea  was  not  substantiated,  however,  by  the  experiments 
made  by  Oddi  and  by  Kettner,^^  who  fed  and  injected  into  animals 
large  quantities  of  the  sodium  salt  of  chondroitin-sulphuric  acid  with- 
out producing  amyloid  changes.  Unpublished  experiments  of  the 
writer  wdth  the  same  material,  as  well  as  with  ground-up  cartilage  and 
with  mucin,  were  equally  unsuccessful.  Likewise  mice  injected  by 
Strada ""  with  the  nucleoprotein  of  pus,  the  so-called  pyin,  or  with 
chrondroitin-sulphuric  acid,  did  not  develop  amyloidosis.  Oestreich  ^ 
injected  cancer  patients  with  chondroitin-sulphuric  acid  for  thera- 
peutic purposes,  but  no  amyloidosis  resulted.     As  it  is  possible   to 

95  Allbutt's  System,  vol.  3,  p.  22.^. 

96Litten  (Verb.  Dent.  Path.  Gesell..  1904  (7),  47)  states  that  thionin  and 
kresyl-violet  are  the  most  specific  stains  for  amyloid,  which  they  color  blue; 
whereas  methyl-violet  stains  red  not  only  amyloid  but  also  mucin,  mast  cell 
granules,  and  tlie  groinul  substance  of  cartilage.  V.  Gieson's  stain  usually  colors 
amvloid  pale  vellow,  and  hvalin  red. 

97  See  Wichmann,  Ziegler's  Reitr.,  1803    (13),  487. 

98  Arch.  exp.  Path.  u.  Pharm.,  1902  (47),  178. 
99Biochem.  Zeit.,  1909    (16),   195. 

1  Zeit.  Krebsforsch..  1911    (11),  44. 


422  RETROGRnSSIVE  CnAXGES 

cause  amj'loidosis  experimentalh'  in  animals,  especially  chickens  and 
rabbits,  by  causing  protracted  suppuration  or  chronic  intoxication 
with  bacterial  filtrates,  these  negative  results  speak  strongly  against 
the  idea  of  a  transportation  of  chondroitin-sulplmric  acid,  but  do  not 
detenuine  it  finally.  They  may  also,  with  propriety',  be  used  in  sup- 
port of  the  statement  of  Hanssen  that  amyloid  does  not  contain  chon- 
droitin-sulphuric  acid.  There  is  usually  much  difficulty  in  producing 
amyloid  experimentally,  for  in  only  a  certain  proportion  of  cases  are 
the  experiments  positive  (in  but  about  one-third  of  Davidsohn's-  100 
trials,  and  many  other  experimenters  have  been  much  less  successful )  .^ 
Davidsohn,  failing  always  to  get  amyloid  experimentally  after  the 
spleen  had  been  removed,  suggests  that  this  organ  (in  which  amyloid 
is  usually  earliest  and  most  abundantly  observed)  produces  an  enzyme, 
which  causes  a  precipitation  of  amyloid  in  the  tissues  from  a  soluble 
precursor  brought  in  the  blood  from  the  site  of  cell  destruction. 
Schmidt  *  considers  it  probable  that  some  enzymatic  action  causes  a 
precipitation  or  coagulation  of  the  substance  in  the  tissue-spaces  or 
lymph-vessels.  Amyloid  is  never  deposited  in  the  cells  themselves,*^ 
and  it  seems  to  be  now  generally  considered  that  the  amyloid  material 
is  infiltrated  in  the  form  of  a  soluble  modification  or  precursor  and 
that  it  is  not  manufactured  in  the  organ  where  it  is  found.  It  is  an 
interesting  fact  that  a  practically  identical  substance  is  formed  in  all 
tissues  and  in  all  species  of  animals,  even  when  the  cause  is  quite  dif- 
ferent. Whether  the  precursors  are  brought  to  the  organ  in  solution, 
or  in  leucocytes,  is  unknown — probably  the  former.  Pollitzer  ^  states 
that  in  various  infections,  especially  coccus  infections,  chondroitin- 
sulphuric  acid  is  excreted  in  the  urine ;  if  this  is  correct  it  has  an 
undoubted  bearing  on  the  genesis  of  amyloidosis.  The  presence  of 
glycothionic  acid  in  pus  ^  is  of  similar  significance.  The  hypothesis 
that  amyloid  is  formed  from  disintegrating  red  corpuscles  is  probably 
incorrect.  Ann'loidosis  is  produced  by  the  most  varied  species  of 
bacteria  and  by  their  toxins,  although  the  staphylococcus  is  usually 
most  effective  in  experimental  work.'^  Neither  is  suppuration  abso- 
lutely essential,  for  injection  of  toxins  alone  {e.  g.,  in  preparing  diph- 
theria antitoxin^),  without  suppuration,  may  produce  amyloidosis,  as 
iilso  frequently  does  syphilis  without  suppuration  and,  less  often, 
many  other  non-suppurative  conditions  {r.  rj.,  tinnors). 

2  Verb.  Deut.  Path.  Gesell.,  1904  (7),  39. 

3  See  Tarchetti,  Deut.  Arch.  klin.  iled..  1903   (75),  526. 

4  Verb.  Deut.  Path.  Cesell.,  1904   (7),  2. 

4a  See  Ebert,  A'ircliow's  Arch.,  1914    (216),  77. 

•-'Deut.  med.  Wocb.,  1912   (3S),  1538. 

<i  .Mandel  and  Levene,  Tiidchciii,  Zcit.,  1907   (4),  7S. 

"  In  a  series  of  experiments  directed  to  ascertain,  if  p()ssihU\  wliich  constituent 
of  pus  niifjlit  he  tlic^  cause  of  amyloid  formation,  1  was  unable  to  secure  amyloid 
by  i)rotracted  intoxication  of  ral>bits  bv  Witte's  "peptom\"  which  consists  chiellv 
of  proteoses   (Trans.  Chicafjo  Path.  Soc.,  1903    (5).  240). 

8  See  Lewis,  Jour.  Med.  Research,  1906   (15),  449. 


UYAfJXK  DEGENERATIOX  423 

Local  amyloid  accumulations  are  of  some  interest  in  considering 
the  genesis  of  the  usual  generalized  form.  They  occur  particularly 
as  small  tumors  in  the  larynx,  bronchi,  nasal  septum,  and  eyelids;  as 
all  these  tissues  are  normally  rich  in  chondroitin-sulphuric  acid,  it 
seems  probable  that  the  amyloid  arises  from  a  local  overproduction  of 
cliondroitin-sulphuric  acid,  which  becomes  bound  with  proteins  in 
.liitu.  This  makes  it  seem  more  probable  that,  in  spite  of  the  lack  of 
positive  experimental  evidence,  general  amyloidosis  is  due  to  liberation 
of  excessive  quantities  of  chondroitin-sulphuric  acid  in  the  sites  of 
tissue  destruction. 

Another  form  of  local  amyloid  is  seen  particularly  in  the  regional 
lymph-glands  of  suppurating  areas;  e.  g.,  the  lumbar  glands  in  verte- 
bral caries,  the  axillary  glands  in  shoulder- joint  suppuration.  This 
local  amyloidosis  is  undoubtedly  due  simply  to  the  fact  that  these 
glands  receive  first,  and  in  largest  amounts,  the  cause,  whatever  it  may 
be,  of  the  amyloid  production. °  Less  readily  explained  are  cases  of 
extensive  amyloidosis  limited  to  the  heart.^° 

Corpora  amylacea  will  be  found  discussed  under  "Concretions" 
(Chap.  xv). 

HYALINE  DEGENEHATION  '^ 

]\Iuch  confusion  concerning  this  condition  may  be  avoided  if  we  ap- 
preciate that  the  term  hyaline  indicates  a  certain  physical  condition, 
which  may  be  exhibited  by  many  substances  of  widely  different  na- 
ture and  origin.  There  is  no  one  chemical  compound,  "hijnlin," 
which,  accumulating  in  cells  or  tissues,  produces  a  hyaline  appear- 
rince.  The  limits  of  the  application  of  the  term  "hyaline  degenera- 
tion," even  to  histological  findings,  is  not  agreed  upon,  but  in  gen- 
eral it  is  used  to  apply  to  clear,  homogeneous,  pathological  substances 
that  possess  a  decided  affinity  for  acid  stains,  such  as  eosin. 
Somewhat  similar  substances,  usually  of  epithelial  origin,  which  do 
not  take  either  acid  or  basic  stains  strongly,  are  usually  called  "col- 
loid." We  may  properly  consider  that  pathological  hyalin  can  be 
divided  into  two  chief  classes  according  to  its  origin:  (1)  connective- 
tissue  hyalin ;  (2)  epithelial  hyalin. 

Connective=tissue  hyalin  is  characterized,  like  amyloid,  by  being 
deposited  in  or  among  the  fibrillar  substance  of  connective  tissues,  and 
not  within  the  cells  themselves,  but  there  are  undoubtedly  several  dif- 
ferent sorts  of  chemical  substances  responsible  for  various  forms  of 

9  Quite  unexplained  is  the  eause  of  the  rarelv  observed  localization  of  amyloid 
in  the  wall  of  the  urinary  bladder.     See  Luckseli    (Verb.  Deut.  path.  Gesell.,  1904 

(7),  34).  Concretions  ofivinp  the  amvloid  reactions  have  been  found  in  the  pelvis 
of  the  kidnev.      (Schmidt,  Cent.  f.  Pathol.,  1912   (23),  86.5.     Mivauchi.  ihid.,  1915 

(26).  289.)  ■ 

10  See  Hecht.  Virchow's  Arch.,  1910  (202),  168. 

11  General  literature,  see  Lubarsch,  Ergeb.  allg.  Path.,  1897  (4),  449. 


424  JiETJiOGRESSJVE  CHAA^GES 

connective-tissue  hyalin.  One  form  is  closely  associated  with  amyloid, 
being-  found  in  organs  showing  amyloid  degeneration,  or  in  other  tis- 
sues in  the  same  body.  In  experimentally  produced  amyloidosis  in 
animals  it  has  been  shown  that  such  a  hyaline  substance  may  appear 
before  the  amyloid,  wliicii  eventually  replaces  it;  hence,  it  has  been 
suggested  that  hyalin  is  a  precursor  of  amyloid.^-  Such  hyalin  differs 
from  true  amyloid  only  in  its  failure  to  give  the  characteristic  stain- 
ing reaction  of  amyloid;  in  all  other  respects,  e.  g.,  cause,  location, 
termination,  it  is  the  same.  As  it  has  been  shown  (see  preceding  sec- 
tion) that  the  staining  properties  of  amyloid  are  vers^  inconstant,  it 
is  probable  that  the  above-described  variety  of  hyalin  is  merely  an  in- 
completely  developed,  or  occasionally  a  reirogressively  altered  amy- 
loid. However,  it  is  probably  not  necessary,  as  some  authors  have 
thought,  that  amyloid  should  always  pass  through  this  hyaline  stage 
in  its  formation. 

Quite  different,  without  doubt,  is  the  form  of  hyalin  observed  in 
.scar  tissue.  This  variety  develops  almost  constantly  in  any  scar-tissue 
after  the  blood-supply  has  been  reduced  to  a  minimum  through  con- 
traction, and  is  seen  characteristically  in  the  corjDora  fibrosa  of  the 
ovary,  fibroid  glomerules  in  chronic  nephritis,  thickened  pleural,  peri- 
cardial, and  episplenitis  scars,  etc.  Such  hyaline  substance  occurs 
independent  of  the  usual  causes  of  amyloid,  affects  only  abnormal 
fibrous  tissue,  never  changes  into  amyloid,  and  is  prone  to  undergo 
calcification — it  surely  has  no  close  chemical  relation  to  the  form  of 
hyalin  that  does  become  amyloid.  Presumably,  it  is  similar  in  na- 
ture to  the  collagen  of  normal  fibrous  tissue  intercellular  substance, 
Avhich  has  undergone  physical  rather  than  chemical  changes  into  a 
homogeneous  hyaline  substance.  For  its  physiological  prototype  it 
has  the  thick  "collagenous"  fibers  of  the  subcutaneous  connective  tis- 
sue. 

Probably  of  quite  different  origin  is  the  hyalin  that  develops  from 
elastic  tissue,  as  seen  best  in  the  thick-walled,  partly  obliterated 
arteries  of  the  senile  spleen;  and  less  characteristically  in  the  early 
stages  of  arteriosclerosis,  since  here  the  preceding  form  of  connective- 
tissue  hyalin  may  also  occur.  Although  arterial  elastic  tissue  is 
related  chemically  to  amyloid,  these  hyaline  vessels  do  not  develop  the 
usual  amyloid  reaction,  but  retain  more  or  less  of  the  specific,  elastic 
tissue  stains.  Presumably  this  form  of  hyalin  is  an  increased  and 
physically  altered  elastin.^^ 

Epithelial  hyalin  occurs  within  the  cells,  and  includes  substances 
of  ])r('siima])ly  widely  diverse  chemical  nature,  from  the  keratin  of 
s(|uamous  epithelium  to  the  snudl  intracellular  hyaline  granules  of 
carcinoma  and  other  degenerating  cells   (Russell's  fuchsin  bodies).^* 

12  Roe  Lubarsch,  Cent.  f.  Pafliol..  1910  (21).  97. 

laSco  Schmidt,  Verh.  Dout.  path.  Cosoll..  1904    (7).  2. 

1*  Literature,  see  llektoen,  Progressive  ^led.,  1899   (ii),  241. 


coLuun  J )i:( n:\JuUATioN  425 

Fiiclisin  l)0(li(\s  are  foniid  also  in  plasma  cells  and,  less  often,  in  other 
cells,  inelu(lin<>-  granulation  tissue;  the  fuchsin  ])odies  of  this  class  are 
believed  by  Brown  ^''  to  be  derived  from  red  corpuscles,  a  view  also 
held  by  Saltykow,  but  not  accepted  by  all  pathologists.'"  Extracellu- 
lar substances  of  hyaline  character,  but  of  unknown  composition,  may 
also  be  i)i"()(luc('(l  by  epitliclium,  c.  g.,  hyaline  casts  in  the  renal  tu- 
bules. 

The  composition  of  none  of  these  forms  of  hyalin  is  known,  except 
that  by  using  microchemical  methods  Unna^^'  has  found  evidence 
that  keratohyalin  consists  of  two  elements,  one  of  acid  character,  ap- 
parently derived  from  the  chromatin,  and  a  basic  substance  resem- 
bling the  globulins. 

Many  other  pathological  materials  of  widely  differing  nature  may, 
under  certain  conditions,  assume  a  hyaline  appearance;  e.  g.,  fibrinous 
exudates  and  thrombi,  degenerated  muscle-fibers  (Zenker's  or  *'waxy" 
degeneration),  tumor-cells  (cylindroma),  etc.  In  all  of  these  the 
chemical  nature  of  the  parent  substance  or  substances  is  probably 
much  less  altered  than  its  physical  appearance,  but  whether  the  change 
is  related  to  the  process  of  protein  coagulation  or  not  is  unknown. 
Occasionally  hyalin,  both  in  epithelium  and  connective-tissue,  takes 
on  a  crystalline  structure  (Freifeld).^^ 

COLLOID  DEGENERATION 

This  term,  also,  has  a  very  indefinite  meaning,  and  is  applied  to 
many  different  conditions  by  various  authors.  Thus,  v.  Reckling- 
hausen includes  under  this  name  amyloid,  epithelial  hyaline,  and  mu- 
coid degeneration.  ^Nlarchand  includes  hyaline  connective-tissue  de- 
generation, and,  also,  as  do  most  other  writers,  the  mucoid  degeneration 
of  carcinoma.  Ziegler  rightly  protests  against  the  inclusion  of  mucin 
under  this  heading,  but  includes  the  corpora  amylacea.  On  account 
of  the  discovery  by  Baumann  of  the  specific  chemical  nature  of  thyroid 
colloid  it  becomes  particularly  unfortunate  that  the  term  "colloid" 
has  such  a  wide  and  uncertain  application.  It  would  seem  that  the 
safest  view  to  take  is  that  the  word  coUold  is  merely  morpJiologically 
and  macroscopicaUii  descriptive  of  certain  products  of  cell  activity  or 
disintegration,  which  have  nothing  in  common  except  the  fact  that 
they  form  a  thick,  glue-like  or  gelatinous,  often  yellowish  or  brownish 
substance.  There  is  no  one  definite  suhstance  colloid,  according  to 
the  usual  usage  of  the  word  in  pathological  literature,  but  many  dif- 
ferent protein  substances  may  assume  the  appearance  to  which  the 

15  Jour.  Exp.  MecL,  1010   (12),  5,33. 

16  See  discussion,  Verh.  Deut.  path.  Gesell.,  lOOS  (12),  26.'S:  :\liintcr,  Virrhow's 
Arch..  1909    (198),  105. 

iGaBerl.  klin.  Woch.,  1914   (51),  598. 

iTZiesler's  Beitr.,  1912  (55),  168;  also  Goodpasture,  Jour.  Med.  Res.,  1917 
(35),  259. 


426  ji'ETh'odh'iussn  j:  vii amies 

name  "colloid"'  is  given.  Looking  at  the  matter  in  this  waj^  we  must 
recognize  as  the  usual  "colloid"  substances,  the  following  chemical 
bodies : 

Thyroid  colloid,  the  physiological  prototype  of  the  group.  This 
consists  of  a  compound  of  globulin  with  an  iodin-eontaining  substance, 
thyroiodin,  the  compound  protein  being  called  bj^  Oswald  iodothyreo- 
globulin.  It  occurs  pathologically  only  in  cystic  and  similar  chauges 
in  the  thyroid  or  accessory  thyroids.  Being  a  specific  product  of  the 
thyroid  (and  perhaps  of  the  hypophysis)  with  definite  physiological 
properties,  it  manifestly  has  only  a  morphological  relation  to  the  other 
forms  of  colloid  found  in  degenerating  tumors,  etc.  In  cysts  of  the 
thyroid,  and  less  often  in  tumors,  there  is  occasionally  found  a  more 
dense  "colloid"  material  of  deeper  color,  the  "caoutchouc  colloid" 
of  the  Germans;  this  seems  to  result  largely  from  transformation  of 
red  corpuscles  in  hemorrhagic  cysts  (Wiget).^*  (The  nature  of  thy- 
roid colloid  is  discussed  more  fully  under  "Diseases  of  the  Thyroid," 
Chap.  XX.) 

Mucin,  when  secreted  in  closed  cavities,  as  in  tumore,  where  it  be- 
comes thickened  by  partial  absorption  of  the  water,  may  take  on  a 
"colloid"  appearance  while  retaining  its  chemical  and  tinctorial  char- 
acteristics. This  is  particularly  observed  in  the  "colloid"  carcinomas 
which  arise  especially  from  the  mucous  membrane  of  the  alimentary 
tract.  This  substance  is,  of  course,  quite  specific  both  in  its  chemical 
luiture  and  its  origin  from  specialized  epithelial  cells,  and  the  process 
should  properly  be  considered  as  a  "mucoid  degeneration." 

Pseudomucin,  which  difi'ers  from  mucin  in  not  being  precipitated 
by  acetic  acid,  is  a  common  component  of  ovarian  cysts,  and  when 
somewhat  concentrated  by  absorption  of  water,  forms  a  "typical 
colloid."  Because  it  is  alkaline,  this  form  of  colloid  tends  to  stain 
rather  with  the  acid  dyes  (eosin,  acid  fuchsin,  etc.),  while  true  mucin 
stains  with  basic  dyes.  Several  varieties  of  pseudomucin  have  been 
described  by  Pfannenstiel,  and  their  properties  will  be  considered  more 
fully  in  the  section  on  "Ovarian  Tumors"  (Chap.  xvii).  The  clear, 
glassy,  yellowish  substance  contained  in  small  cavities  of  ovarian  tu- 
mors, which  is  usually  called  ' '  colloid, ' '  consists  of  nearly  pure  pseudo- 
mucin. All  these  substances  yield  a  reducing  substance  on  boiling 
with  acids,  which  is  a  nitrogen-containing  body,  f/Jucosamin^^ 

Simple  proteins  {e.  g.,  serum-globulin,  serum-albumin,  nucleo- 
albumin,  etc.)  may,  when  in  solution  in  closed  cavities,  become  con- 
centrated through  absoi*ption  of  water  until  they  produce  the  physical 
appearance  of  "colloid."  Probably  the  colloid  contents  of  dilated 
renal  tubules,  cavities  in  various  mesoblastic  tumors,  etc.,  are  pro- 
duced in  this  way. 

isVirchow's  Arcli.,  1000   (IS;")),  410:  von  Simior,  ibid.,  lOl.j    (210),  2711. 
10  Ziinpcrle.  Miindi.  mod.  Wodi.,  10(10   (47).  414. 


MUCOID  DEaEXEh'ATWy  ^27 


MUCOID  DEGENERATION 

ATnoin    in  its  typical  form,  is  a  compound  protein,  consisting  of  a 
:\Iucm,  in  Its  IH  ica  '     ^    ^^^^i^      carbohydrate,  rjlucosamm. 

S:;:r:t;r1>oerwir:crds    mucin  ;Ma.  .  substanee  reducing 
Sno-'s  so  ution.     Mucin  is  acid  in  reaction,  probably  because  of 

:  thZc'dvL*  It  r;rdily  dissolved  in  very  weak  alkaline  solu^ 
n  s  is  prec Stated  br  acetic  acid,  and  its  physical  properties  when 
n  inrirarfc  ui.e  characteristic.  The  t;""™-".  >;---;  ^tl 
Wv  covers  a  number  of  related  but  distinct  bodies.  Some,  sucli  as  tne 
"fc.  r^c^rare  readily  distinguished  by  not  b^ing  preeipi  a  -  b^ 
acetic  acid  and  bv  being  alkaline  in  reaction:  others  jield  ieducin„ 
suWances  without  previous  decomposition  with  acids  (paramucm  ; 
while  even  Tmong  tile  "true"  mucins  certain  differences  in  solubility 

'tnThe  mammalian  body  we  find  mucin  <"=.<;"■■>;!"=  '■;,f™2f'ta  tt 
calitics:  (1)  as  a  product  of  secretion  of  ePf ^^^  Is  (2^  m  the 
interstices  of  connective  tissue,  especially  of  t  ndo.  .  (^^^  J^^J 
Hanee  of  svnovial  fluid  to  mucin  is  more  physica  than  ™emical.) 
Thpre  is  a  so  evidence  that  mucin  or  a  related  body  constitutes  the 
cement  suSance  between  all  the  bodycells.  Corresponding  to  these 
Zc^iief  sources  of  mucin  we  And  mucoid  degeneration  occnrring  as 
;^stincrprocesses  in  mucous  membranes  (or  tissues  derived  therefrom) 

'^Eprthl'ilLT  M^cin!-!  epithelial  mucin  represents  a  distinct  prod- 
uct ospcialized  cells,  it  is  questionable  if  the  ordinary  app. cation 
o    th    tenndegeneration  in  the  sense  of  the  conversion  of  eell-proto- 
nlasr^    nto  mucin,  is  correct.     Certainly  the  mucin  formation  of  ca_ 
t'arhal    nflaliation  is  merely  an  excess  of  a  -"->  --  '^j^^;^ 
tbP  deo-enerative  changes  that  may  be  present  m  the  epithelial  cells  are 
nroducedbv  the  cause  of  the  inflammation,  and  are  not  dependent  upon 
mucirfornn.  ion.     Even  in  the  extreme  example  of  mucoid  degenera- 
tion" en  i^  carcinomas  derived  from  mucous  membranes  (the  so-c<dU^ 
••?ollok""ancers"),  the  epithelial  degeneration  is  not  necessanh   to 
he    nterme"ed  as  a  conversion  of  cell-eytopla,sm  into  mucin,  but  is 
largely  due  to  the  pressure  of  secreted  mucin  upon  the  cells  within 
,.For  special  consideration  «e  Cutter  and  Oies,  .^mer.  .Tour,  rli.vsiol..   IWl 

""■  '^?-  ,      ,„  ■.        „    P.ll,     ion    (14)     23)    savs  that  the  long  controversy 

so.  Sohadc    IZeit.  exp.  Patli..   l-HJ    ' ','•  -;J'    ,|-'     ,,„„„crtive  tissue  is  settled 

eoneernins  the  intercellular  '"''»;»„""  „°/""^™„k?"rt"S    p.  321).  who  found 

trpr„™irr;l5':ttS:rand"f:,dr|ucin  is  indicate.,  ,.  ^ 
logical  inter-reactions  (Elliott,  Jour.  Infect.  Dis.,  1014   (la),  oUi). 


428  RETROdUEiiiiiyE  CUAXGES 

tlie  confined  spaces  of  the  tumor.  Tlie  mucin  in  these  forms  of  mucoid 
(k^gencration  is  cliemically  tlie  same  as  the  normal  mucin  coming  from 
the  same  source,  but  mixed  with  larger  or  smaller  quantities  of  other 
proteins  derived  from  cell  degeneration  or  from  vascular  exudates, 
and  we  do  not  yet  know  certainly  the  chemical  character  of  the  secre- 
tion  of  iioi'mal  mucous  monibranos.-'"'  (The  stringy,  mucin-like  sub- 
stance seen  in  some  purulent  exudates  is  probably  composed  largely  of 
nueleoproteins  and  nucleo-albumins  derived  from  the  degenerating 
leucocytes,  and  is  not  true  mucin.) 

Connect! ve=tissue  Mucin. — Excessive  formation  of  connective-tis- 
sue mucin  is  observed  most  characteristically  in  myxedema  {q.  v.), 
but  may  also  occur  in  connective  tissues  that  are  poorly  nourished  or 
otherwise  slightly  injured ;  it  is  seen  particularly  in  the  connective 
tissues  surrounding  the  epithelial  elements  in  adenomas  and  carcino- 
mas. In  the  walls  of  large  blood  vessels  there  is  a  mucoid  connective 
tissue,  rich  in  mucin,  which  may  be  increased  in  arterio-sclerosis 
(Bjorling).-^  Connective-tissue  tumors  (myxosarcoma,  myxofibroma, 
or  myxoma)  may  also  show  a  great  quantity  of  mucinous  intercellular 
substance,  but  many  of  the  so-called  myxomas  are  in  reality  merely 
edematous  fibromas  or  polypoid  tumors,  in  which  the  resemblance  to 
true  myxoma  is  largely  structural  rather  than  chemical.  This  form 
of  mucoid  degeneration  seems  to  be  merely  a  reversion  to  the  fetal  type 
of  connective  tissue,  which  is  characterized,  as  in  the  umbilical  cord,  bj^ 
an  excessive  accumulation  of  a  mucin-containing  fluid  intercellular 
substance,  and  a  paucity  of  collagenous  fibrillar  structure.  Appar- 
ently, when  connective  tissue  reverts  to  an  embryonal  type,  either 
from  intrinsic  causes  (tumor  formation),  or  when  the  nourishment 
is  insufificient,  or  possibly  when  the  normal  stimulus  to  cell  growth  is 
absent  (myxedema),  the  mucoid  characteristics  of  fetal  tissue  reappear. 

The  presence  of  mucin  in  the  tissues  seems  to  cause  no  reaction, 
and  its  absorption  causes  no  harm.  Rabbits  that  I  injected  with 
large  quantities  of  pure  tendon  mucin  almost  daily  for  two  to  four 
months,  showed  absolutely  no  deleterious  effects,  either  locally  or  con- 
stitutionally. Some  of  the  French  authors  --  claim  that  mucin  pos- 
sesses a  slight  bactei'icidal  power.  On  the  other  hand,  Rettger -^  and 
others  have  found  an  apparently  typical  mucin  produced  by  certain 
varieties  of  bacteria. 

GLYCOGEN  IN  PATHOLOGICAL  PROCESSES  -* 

It  seems  probable  that  all,  or  nearly  all,  cells  contain  larger  or 
smaller  quantities  of  glj^cogen,  but  it  may  be  insufficient  in  amount 

20b  See  Lopez-Suaroz,  Bioehcm.  Zoit.,  101.3   (56),  107. 

21  Vircliovv's  Arohiv.,  ]!»]]    (2().")),  71. 

22  Arloinsr.  Compt.  Tlond.  Soo.  Biol..  1002    {TA),  .100.  and   1001    (53).  1117. 

23  .Tour.  :\Ic(l.  Bcsoarcli,  100.3    (10),  101. 

24  Biblidfrrapliv  by  fliorko,  Ziosrlor's  Boiir..  1005  (.37).  502,  and  Ergcbnisse 
Pathol.,  1007,  Xi(j)",  871;   Klosfadt,  ibid.,  1011    (XV(,),  .349. 


aiACOdhX  J\   I'A'lllolJJGICAL  PROCEHHEH  429 

to  be  detected  citliei-  7Hieroseoi)ically  oi"  clu'inically.  Glycogen  seems 
to  be  formed  within  tlie  cells  from  the  sugar  of  the  blood,  through  a 
process  of  dehydration  and  polymerization,  and  to  be  reconverted 
whenever  necessary  into  sugar,  by  a  reverse  process  of  hydrolysis.  It 
is  quite  possible  that  both  of  these  processes  represent  merely  the 
reversible  action  of  an  intracellular  enzyme,  but  this  has  not  been 
established.  We  do  know,  however,  that  soon  after  death  the  intra- 
cellular glycogen  is  rapidly  converted  into  dextrose.-^ 

Properties  of  Glycogen. — Clycofxen  is  fieciiu'iitly  called  an  "animal  starch," 
liavinjj  the  sanio  uoiicial  composition  as  tho  starches  ( r„H,„0,-, )  a?,  and  apparently, 
like  the  starches,  it  represents  a  relatively  insoluble  rcstinfr  staf,'e  of  sufjar  in 
the  course  of  nietaholism.  It  is  readily  solulilc  in  water,  formintf  an  opalescent, 
colloidal  solution,  and,  therefore,  lias  no  cITect  on  osmotic  pressure,  and  it  is  not 
diffusible. 2C  Because  of  its  soluI)ility  and  the  rapidity  witli  wliicli  postmortem 
<"hange  to  dextrose  occurs,  specimens  that  are  to  be  examined  microscopically  for 
glycogen  must  l)e  hardened  while  very  fresh  in  strontj  alcohol,  in  which  srlycojren 
is  insoluble. 2"  One  of  the  most  characteristic  reactions  is  the  port-wine  color 
^iven  by  glycofjen  when  treated  with  iodin ;  this  reaction  may  be  applied  micro- 
scopically, solution  of  the  glycogen  being  avoided  by  having  the  iodin  dissolved  in 
a  solution  of  gum  arable  or  in  glycerol.  Salivary  ptyalin  rapidly  converts  gly- 
cogen into  glucose,  and  this  reaction  may  also  be  used  microscopically  to  prove 
that  suspected  granules  are  glycogen.  However,  failure  to  find  glycogen  micro- 
chemically  does  not  always  mean  its  absence  from  a  tissue. ^s 

PHYSIOLOGICAL  OCCURRENCE 

According  to  Gierke,  the  normal  glycogen  of  cells  resembles  fat 
in  that  part  of  it  disappears  during  starvation,  while  the  rest  cannot 
be  removed  in  this  way  and  probably  is  something  more  than  a  re- 
serve food-stutf.  In  distribution  glycogen  somewhat  resembles  fat, 
being  abundant  in  the  liver  ~^  and  muscles,  but  Gierke  considers 
that  the  microscopic  evidence  of  the  quantity  of  glycogen  present  in 
the  cell  agrees  better  with  the  results  of  actual  chemical  analysis  than 
is  the  case  with  fat.  Rusk,''"  however,  tinds  only  a  general  agreement, 
wdth  marked  exceptions.  Neither  iodin  nor  Best's  carmin  stain  are 
absolutely  specific  for  glycogen,  but  Gierke  believes  that  we  may 
safely  consider  a  substance  as  glycogen  when  it  is  homogeneous, 
rather  easily  soluble  in  water  and  more  so  in  saliva,  gives  the  usual 
iodin  reaction,  and  stains  bright  red  with  Best's  carmin  solution. ^^ 
With  these  controls,  the  microscopic  findings  were  found  to  agree 
closely  with  the  results  of  direct  chemical  analysis,  and  glycogen  was 
found  microscopically  visible  in  muscle,  liver,  lung,  heart,  uterus,  and 

25  Literature  concerning  phvsiolo2"\'  of  glycogen  by  Pfliiirer.  Pfliifrer's  Arch., 
1903   (96),  39S:  and  Cremer.  Frireb.'der  Phvsiol..  1902   (1,  Abt.  1),  Sn.3. 

26  See  Gatin-Oru/ewska,  Pfliiger's  Arch..  1904    (10.3),  282. 

27  According  to  TTelman  (Cent.  f.  inn.  ]\fed..  1902  (2.3),  1017),  glycogen  may  be 
found  in  specimens  preserved  in  alcohol  as  long  as  fifteen  years. 

28Bleibtreu  and  Kato,  Pfliiger's  Arch.,  1909   (127).  118.' 

29  In   the  livers  of  two  executed   criminals   Oarnier    (Compt.   Pend.   Soc.   Biol., 
1906    (60),  12.5)    found  respectively  4  per  cent,  and  2.79  per  cent,  of  glycogen. 
3oUniy.  of  California  Publ.,  Pathol.,  1912   (2),  83. 
31  Concerning  staining  methods  see  Klestadt,  Joe.  cit. 


430  RETROGRESSIVE  CHANGES 

skin  (but  not  in  the  brain,  where  it  maj^  be  demonstrated  chemically 
in  minute  quantities). 

Glycogen  is  commonly  said  to  be  especially  abundant  in  fetal  tis- 
sues, but  it  is  not  present  in  all  fetal  cells,^-  nor  is  it  always  most 
abundant  in  the  most  rapidly  growing  tissues.  Althoug-h  both  fat 
and  glycogen  are  quite  abundant  in  fetal  muscle  and  liver  tissues, 
the  liver  of  early  embryos  does  not  contain  either.^^  Invertebrates 
and  the  lower  vertebrates  have  more  than  the  higher  forms.  In  mam- 
malian adults  the  liver  and  muscle  contain  the  most  glycogen,  carti- 
lage standing  next,  and  it  is  also  present  in  squamous  epithelium 
(particularly  the  middle  layers),  especially  that  of  the  vagina  (Wieg- 
mann),  but  not  in  slightly  stratified  (cornea),  transitional,  or  cylin- 
drical epithelium.  Normal  human  kidneys  do  not  seem  to  show  gly- 
cogen, but  it  may  be  present  in  the  kidneys  of  mice,  rabbits,  and 
cats.  There  is  considerable  in  the  heart  muscle.^*  Gh^cogen  is  most 
abundant  in  the  utenis  at  the  time  of  child-])irth,  and  is  abundant  in 
the  placenta ;  but  it  is  also  present  in  the  uterus  and  tubes  independent 
of  pregnancy.^*''  After  pancreas  extirpation,  Fichera  ^^  observed  a 
disappearance  of  all  visible  glycogen,  except  a  little  in  the  cartilage 
and  stratified  epithelium ;  hence  he  considers  the  glycogen-content  as  a 
function  of  cell  nourishment.  Fat  and  glycogen  often  occur  together, 
although  one  may  be  present  without  the  other  (Gierke).  Presuma- 
bly the  failure  to  find  glj^cogen  in  certain  cells  depends  rather  on  a 
failure  of  technic  than  on  a  total  absence  of  glycogen. 

There  has  been  some  diversity  of  opinion  as  to  whether  glycogen 
occurs  as  granules  in  the  living  cell,  or  whether  the  granules  are 
formed  from  a  homogeneous  substance  by  hardening  fluids.  In  view 
of  the  clear-cut,  definite  spaces  it  may  leave  in  cells  when  dissolved 
out,  glycogen  probably  occure  as  granules,  especially  when  present 
in  abnormally  large  quantities.  The  studies  of  Arnold  have  shown 
that  in  many  cells  the  glycogen  takes  on  a  definite  structure,  in  close 
relation  to  the  plasmosomes.  It  has  been  suggested  that  the  intra- 
epithelial hyaline  bodies  (Russell's  fuchsin  bodies)  are  glycogenic, 
w^hich  idea  is  probably  not  correct.  Habershon  and  others  have  sug- 
gested that  eosinophile  granules  are  either  glycogen  or  related  to  it. 
The  presence  of  glycogen  in  the  cells  seems  to  cause  no  injury  to  the 
cytoplasm,  and  if  it  again  disappears,  the  cells  become  quite  normal.^" 

32Spp  Glinko.  Biol.  Zoit..  Moskau.  1911    (2),  1. 

33  AdainofT  (Zeit.  f.  Biol..  1005  (46),  288)  contests  the  idea  that  the  amount 
of  frlycofren  is  in  direct  relation  to  <rro\vth  enortry;  see  also  !Mendel  and  Leaven- 
worth (Amer.  Jour.  Physiol.,  1007  (20),  117),  who  found  no  particular  abundance 
in  the  tissues  of  the  fetal  pip. 

3+Berl)linjrer,  Ziealer's  Beitr.,  1012   (".3),  155. 

34a  :McAllister,  Jour.  Obs.  Clvn.  Brit.  Emp.,  101.3    (34),  01. 

sr.  Ziepler's  Beitr..  1004    (3fi).  273.  literature. 

30  Yet  Teissier  (Compt.  Bend.  Foe.  Biol..  1000  (52),  700)  I)elieves  the  amount 
normally  present  in  the  liver  is  strongly  bactericidal,  and  in  a  hater  publication 
{Hid.,   1902    (54),   1098)    considers  that  it  is  toxic  to  liver-cells.     Wendelstadt 


(ILYCOdEX  I\   I'ATIIOIAXIICAL  J'h'OCES.SES  431 

Even  the  nuclei  may  contain  granules  of  glycogen  without  evident 
permanent  injury. 

PATHOLOGICAL  OCCURRENCE 

According  to  the  results  obtained  by  Fichera  and  Gierke,  it  seems 
probable  that  glycogen  accumulation  is  produced  under  the  same 
conditions  as  are  fatty  changes,  i.  e.,  when  oxidation  is  locally  or 
generally  impaired.  Fat  and  glycogen  are,  therefore,  often  found 
together  in  the  margins  of  infarcts  and  of  tubercles,  in  passive  con- 
gestion of  the  liver,  and  in  heart  muscle  with  fatty  changes  due  to 
severe  anemia.  The  glycogen,  being  more  labile,  seems  to  disappear 
early  when  the  cells  become  necrotic,  and  hence  glycogen  is  not  pres- 
ent in  older  necrotic  areas  where  the  fat  still  persists.  (This  proba- 
bly accounts  for  the  frequently  repeated  statement  that  glycogen  and 
fat  do  not  occur  together.)  Whether  the  glycogen  can  be  trans- 
formed into  fat,  perhaps  forming  an  intermediary  stage  in  a  trans- 
formation of  protein  into  fat,  has  not  been  determined,  but  there 
seems  to  be  little  doubt  that  it  is  infiltrated  from  outside  the  cell, 
and  not  formed  directly  from  degenerated  protein.  It  seems  to  be 
deposited  only  in  cells  that  are  still  living,  although  it  can  become 
split  up  in  dead  cells.  All  cells,  but  especially  muscle-cells  and  leu- 
cocytes, seem  able  to  lay  up  glycogen  in  visible  amounts  under  cer- 
tain conditions.  In  inflamed  areas  glycogen  is  found  in  both  tissue- 
cells  and  leucocytes,  but  not  in  cells  showing  nuclear  degeneration 
(Best,  Gierke).  In  pneumonia  the  leucocytes  of  the  exudate,  and 
to  a  less  extent  the  alveolar  epithelium,  contain  glycogen  as  well  as 
fat.  In  tubercles  glycogen  is  found  in  the  cells  which  contain  ba- 
cilli, and  it  is  generally  present  in  the  epitheloid  cells,  rarely  in  giant 
cells,  not  at  all  in  lymphoid  cells  or  in  the  necrotic  elements  (De- 
vaux).  Liver  glycogen  is  altered  most  in  poisoning,  being  reduced 
by  phosphorus,  arsenic,  chloroform,  IlgCU,  and  many  other  poisons; 
the  amount  is  reduced  when  death  from  any  cause  is  slow,  or  when 
putrefaction  has  occurred,  but  it  is  increased  in  carbon  monoxide 
poisoning  (Massari).^^  In  rabbits,  at  least,  it  is  deposited  in  the  liver 
first  about  the  central  vein,  and  in  fasting  animals  it  disappears  first 
from  the  periphery-."** 

Glycogen  in  Tumors. — Glycogen  has  been  observed  frequently  in 
tumors.  Brault  believed  the  quantity  an  index  of  rate  of  growth,  on 
the  principle  that  glycogen  appears  most  abundantly  in  embryonal 
tissues,  and  therefore  in  tumors  the  amount  of  glycogen  should  agree 
with  the  degree  to  which  the  cells  have  gone  back  to  the  embryonic 
type.  Lubarsch  considered  that  only  tissues  normally  containing 
glycogen  give  rise  to  glycogen-containing  tumors.     Gierke  could  cor- 

(Cent.  f.  Bact.,  Abt.  1,  100,3   (34),  831)    found  that  iiiulpr  certain  conditions  gly- 
cogen impedes  hemolysis  by  normal  serum. 

STGax.  degli  Ospedali,   1006    (27),  .537. 

sslshimori,  Biochem.  Zeit.,  1913   (48),  332. 


432  RETROGRESSIVE  CHAXGES 

roborate  neitlier  of  these  ideas,  and  considers  that  gl3^cogen  appears  in 
tumors  under  exactly  the  same  conditions  in  which  it  appears  in  other 
tissues;  i.  e.,  when  cell  nutrition  and  oxidation  are  impaired.  Ap- 
parently, however,  hoth  the  emhryonic  origin  and  local  retrogressive 
changes  determine  the  deposition  of  glycogen  in  tumors.  Glycogen 
is  particularly  abundant  in  squamous  epithelium  of  epitheliomas  that 
have  gone  on  to  homification ;  ^'^  in  testicular  tumors,  hyperneph- 
romas, parathyroid  tumors  (Langhans),*"  endotheliomas,  chondromas, 
and  myomas,  and  it  also  occurs  in  the  connective  tissues  surrounding 
tumors.  Of  1544  tumors  of  all  sorts  examined  by  Lubarsch,^^  447 
(or  29  per  cent.)  contained  glycogen  microscopically;  fibromas,  oste- 
omas, gliomas,  hemangiomas  were  always  free  from  glycogen;  and 
lipomas  and  lymphangiomas  nearly  always.  Adenomas  are  almost 
equally  free  from  glycogen  (two  positive  in  260  specimens),  while  it 
was  constant  in  teratomas,  rhabdomyomas,  hypernephromas,  and 
chorioepitheliomas.  Fifty  and  seven-tenths  per  cent,  of  the  sarcomas 
and  43.6  per  cent,  of  the  carcinomas  show  glycogen,  most  abun- 
dant in  squamous-cell  epitheliomas ;  columnar-celled  carcinomas  con- 
tain glycogen  much  less  often,  and  it  is  always  absent  in  "colloid 
cancers. ' ' 

Animal  parasites,  in  common  with  other  invertebrates,  usually  show 
abundant  quantities  of  glycogen.'*-  It  has  been  found  in  protozoa,  as 
well  as  in  all  varieties  of  intestinal  worms.  According  to  Barfurth, 
nematodes  in  glycogen-free  animals  may  contain  glycogen.  The  gly- 
cogen is  found  chiefly  in  the  connective  tissues  of  the  intestinal  para- 
sites, but  in  some  of  the  nematodes  it  occurs  chiefly  in  the  sexual 
organs  and  muscle-cells.  The  walls  of  the  hydatid  cysts  contain  much 
glycogen,  which  is,  perhaps,  related  to  the  usual  presence  of  sugar 
in  their  contents.  If  Habershon's  contention  is  correct,  that  eosino- 
phile  granules  are  related  to  glycogen,  we  may  have  here  an  expla- 
nation of  the  occurrence  of  eosinophilia  in  infection  with  animal  para- 
sites.    (See  also  "Animal  Parasites,"  Chap,  v.) 

Glycogen  in  Leucocytes. — The  occurrence  of  glycogen  in  the  blood 
has  aroused  nuicli  interest,  particularly  in  relation  to  its  diagnostic 
value.  INIany  leucocytes  contain  granules  that  stain  with  iodin,  and 
although  it  is  possible  that  these  are  not  all  granules  of  glycogen, 
yet,  for  the  most  part,  they  probably  represent  this  substance  in 
excessive  quantities.  The  granules  are  observed  chiefly  in  the  poly- 
morphoinicloar  neutrophiles,  but  also  in  large  and  small  mononuclear 
cells  and  cosinophiles.     Occasional  granules  arc  also  found  free    (or 

■''!>  Tn  mouse  tvimora  TTaaland  found  fflycofren  only  in  squamous  coll  caroinoma, 
and  in  the  connective  tissue  aurroundinfr  other  tumors  (Jour.  Path,  and  Tiact., 
H»OH    (12).  4.3!)). 

40Virchow's  Arch.,   1007    flSn),   1:58. 

41  Virchow's  Arch.,  1900   (183).  188. 

•*-  Elaborate  treatise  on  occurrence  of  <rlyco<ren  in  lower  aniinals  by  I'arfurth, 
Arch,  niikros.  .Xnat.,  1885  (2.5).  200;  also  r>usch.  Arch,  internat.  phvsiol..  1005 
(3),  49;  Brault  and  Loeper,  Jour.  Phys.  et  Path.  G(n\.,  1004   (6),  205  and  720. 


dLYVOUKS  l\    rATII()L()(;l('AL   PltOCKHtiES  433 

perhaps  contained  in  blood-platelets)  in  all  blood,  wliether  normal 
or  patliolo<^ieal.  '■'■  nirscliber<?  ^^  states  that  normal  animals  of  all 
species  have  leucocytes  jiivin*^  an  iodin  reaction  for  glycogen  if  pi-oper 
techuic  is  used,  but  which  is  not  obtained  by  the  ordinary  iodin-gum 
solution  method  unless  the  glycogen  is  rendered  abnormally  insolu- 
ble by  toxic  injury;  this  is  an  explanation  for  the  relationship  of 
i()d()j)liilia  and  infections.  Accordino'  to  Wolff- Eisner  the  leucocytes 
in  myeloid  leukemia  contain  no  glycogen  granules.  It  does  not  seem 
to  be  settled  whether  the  glycogen  is  taken  on  by  the  leucocytes  at 
the  place  of  pathological  lesion,  or  in  the  bone-marrow  under  the  in- 
fluence of  circulating  poisons,  or  both.  TTabershon  states  that  from 
1  to  16  per  cent,  of  all  leucocytes  normally  contain  glycogen  granules, 
and  AVolff  believes  that  the  glycogen  seen  in  leucocytes  represents 
normal  glycogen  made  insoluble  through  injury. 

Locke  gives  the  occurrence  of  this  abnormal  iodin  staining  of  the 
leucocytes  (termed  iodophilia)  as  follow'S:  "Septic  conditions  of  all 
kinds,  including  septicemia,  abscesses,  and  local  sepsis  (except  in  the 
earliest  stages),  appendicitis  accompanied  by  abscess  formation  or  per- 
itonitis, general  peritonitis,  empyema,  pneumonia,  pyonephrosis,  sal- 
pingitis with  severe  inflammation  or  abscess  formation,  tonsillitis, 
gonorrheal  arthritis,  and  hernia  or  acute  intestinal  obstruction  where 
the  bowel  has  become  gangrenous,  have  invariably  given  a  positive 
iodophilia,  and  by  its  absence  all  these  cases  can  be  ruled  out  in  diag- 
nosis. In  other  words,  no  septic  condition  of  any  severity  can  be 
present  without  a  positive  reaction.  Furthermore,  the  disappearance 
of  the  glycogen  granules  in  the  leucocytes  in  from  twenty-four  to 
forty-eight  hours  following  crisis  with  frank  resolution  in  pneumonia, 
and  the  thorough  drainage  of  pus  in  septic  cases,  is  of  considerable 
importance."  Clinical  experience,  however,  seems  not  to  have  ac- 
corded any  constant  significance  to  iodophilia.''^ 

In  exudates  glycogen  is  found  in  the  leucocytes  as  long  as  they 
retain  their  vitality,  but  disappears  soon  after  retrogressive  changes 
begin ;  hence  it  is  not  usually  present  in  sterile  pus.  Loeper  *°  made 
quantitative  estimates  of  the  glycogen  in  exudates,  finding  from 
0.59-0.62  gram  per  liter  in  cellular  pneumococcus  pleural  effusi6n, 
0.25  gm.  in  cellular  tuberculous  effusion,  but  only  traces  in  serous 
tuberculous  efiPusion  and  in  an  old  tuberculous  pyothorax.  A  pneu- 
monic lung  contained  0.85  gm.  of  glycogen  per  kilo,  and  traces  were 
found  in  pneumonic  sputum  and  in  the  contents  of  tuberculous  cavi- 
ties.    It  is  very  abundant  in  tuberculous  sputum,  as  much  as  2  to  3  per 

43  Literature — Locke  and  Cabot,  Jour.  Med.  Researeh,  1002  (7),  25:  Locke, 
Boston  Med.  and  Surg.  Jour.,  1002  (147).  289:  Reich.  Beitr.  klin.  Chir.,  1!)04  (42), 
277;  Kiittner,  Arch.  klin.  Chir..  1004  (73),  438;  CuHand,  Brit.  :\red.  Jour..  1004 
(i),  880;  Ilahershon,  Jour.  Path,  and  Bact.,  lOOG  (11),  05;  Woin",  Zeit.  klin. 
Med.,  1004   (51),  407. 

44Virchow's  Arch.,  1008    (194),  307. 

45  See  Bernicot.  Jour.  Path,  and  Bact.,  1006    (11),  304. 

46  Arch.  Med.  Exp.,  1902   (14),  576. 

28 


434  RETROGRESSIVE  CHAyCES 

cent,  in  advanced  stages,  but  absent  in  bronchial  catarrh ;  in  pneu- 
monia .05  per  cent,  was  found,  in  putrid  bronchitis  0.25  per  cent. 
(Pozzilli).  When  glvcouen  solution  (1  per  cent.)  is  injected  into  the 
peritoneal  cavity,  the  endothelial  cells  and  invading  leucocytes  become 
loaded  with  glycogen  granules. 

Glycogenic  Infiltration  in  Diabetes. —  It  is  in  diabetes,  however^ 
that  tiie  iiKist  marked  acnimulatious  of  glycogen  are  found,  the  gran- 
ules frequently  fusing  in  the  cells  into  droplets  larger  than  the  nu- 
cleus ;  when  dissolved  out  in  ordinary-  microscopic  preparations,  the 
clear  round  space  left  is  exactly  like  the  space  left  by  a  fat-droplet, 
except  that  the  margins  show  a  tendency  to  take  the  basic  stain  for 
some  unknown  reason.  In  even  the  most  extreme  cases,  however, 
the  nucleus  is  well  preserved,  although  it,  too,  may  contain  large 
masses  of  glycogen,  in  which  case  there  is  no  glycogen  in  the  cyto- 
plasm.*^ Glycogen  is  found  particularly  in  the  epithelium  of  Henle's 
tubules,*'^  in  heart  muscle,  and  in  the  leucocytes,  whereas  it  is  greatly 
diminished  in  the  normal  storehouses  of  glycogen,  the  liver  and  mus- 
cles. Fiitterer  describes  masses  of  glycogen  in  the  cerebral  capil- 
laries, resembling  an  embolic  process;  it  is  also  present  in  the  tissues 
of  the  eye.**'  Sandmeyer  analyzed  the  organs  for  glycogen  in  a  case 
of  diabetes,  finding  the  following  amounts  in  percentage  of  organ 
weight:  liver,  0.613;  kidneys,  0.1158;  lungs,  0.0442;  spleen,  0.07. 
Experimental  diabetes  (pancreas  extirpation)  produces  a  marked  glj'- 
eogenic  infiltration. 

47  Askanazy  and  Hiibschmann,  Cent.  f.  Path.,  1907    (18),  641. 

48  See  Fahf,  Cent.  f.  Path.,  1911    (22),  94.5. 

49  Shimagav/ora,  Klin.  Monatsbl.  Augenheilk.,  1911    (12),  682. 


CHAPTER   XV 

CALCIFICATION,  CONCRETIONS,  AND 
INCRUSTATIONS 

CALCIFICATION ' 

Pathological  calcitication  occurs  in  two  forms:  one  is  a  precipita- 
tion of  calcium  in  secretions  and  excretions  of  the  body;  the  other 
is  the  deposition  of  calcium  salts  in  the  tissues  themselves.  The 
former,  which  includes  not  only  concretions  in  general,  but  probably 
also  the  deposition  of  calcium  salts  in  the  cells  and  tubules  of  the 
kidney,-  both  in  disease  and  in  experimental  calcification  after  cer- 
tain poisonings,  is  readily  enough  explained  in  most  instances  by  rec- 
ognizable alterations  in  the  composition  of  the  secretions,  which  lead 
to  simple  chemical  precipitations.  With  this  form  w^e  shall  deal  in 
the  subsequent  consideration  of  concretions,  but,  in  referring  to  calci- 
fication, shall  indicate  only  depositions  from  the  blood  directly  into  the 
tissues. 

Relation  of  Calcification  to  Ossification. — In  nonnal  ossification 
we  have  to  deal  with  the  accumulation  of  lime  salts  within  the  stroma 
or  cells  of  a  tissue  that  has  usually  undergone  certain  preparatory 
changes  in  the  way  of  formation  of  a  more  or  less  homogeneous 
ground  substance,  but  has  not  suffered  a  total  loss  of  vitality,  al- 
though vitality  is  possibly  decreased.  Pathological  calcification  is 
similar,  in  so  far  as  we  have  to  deal  Math  deposition  of  quite  the  same 
salts  in  tissues  that  have  suffered  either  total  or  partial  loss  of  vital- 
ity, and  which  ver\^  frequently  indeed  are  hyaline.  Wliat  appear  to 
be  essential  differences  are  these:  (1)  In  calcification  the  lime  salts 
always  remain  in  clumps  and  masses,  often  fusing  to  greater  or  less 
degree,  but  never  with  the  diffuse  even  permeation  of  tissue  seen  in 
ossification.  (2)  All  the  cells  within  a  calcified  area,  if  not  dead 
at  the  beginning  of  the  process,  eventually  disappear  for  the  most 
part,  and  we  have  sooner  or  later  a  perfect!}-  inert  mass,  practically 
a  foreign  body,  instead  of  a  specialized  tissue  as  in  ossification.  (3) 
Ossification  is  accomplished  only  in  varieties  of  connective  tissue, 
but  calcification  may  involve  any  sort  of  cell  or  tissue  provided  it 
is   degenerated   sufficiently.     Furthermore,   any   area  of   calcification 

1  Literature  and  r#siim#:  Pfaundlcr,  Jahrb.  f.  Kinderheilk..  1904  (00),  123; 
Wells,  Jour.  [Nfed.  Research,  1006  (14),  491,  and  Arch.  Int.  Med..  1011  (7),  721; 
Hofmeister.  Ergebnisse  Physiol.,  1910  (9),  429;  Schultze,  Ergebnisse  Pathol., 
1910,  XIV   (2),  706. 

2  See  Wells,  Holmes  and  Henry,  Jour.  Med.  Research,  1911   (25),  373. 

435 


436 


CALCIFICATIOX,    COXCRETIOXS,    AND    INCRUSTATIOyS 


is  likely  to  be  replaced  by  bone,  no  matter  what  tissue  may  be  in- 
volved ;  apparently  the  presence  of  calcium  salt  deposits  in  any  part 
of  the  body  can  stimulate  the  connective  tissues  to  form  bone,  but  in 
the  absence  of  calcium  salts  even  the  cells  which  are  normally  osteo- 
genic will  not  form  bone. 

Composition  of  the  Deposits  in  Calcification.-^ — The  composition 
of  the  inoryaiiic  salts  in  ealeilied  areas  in  the  body  seems  to  be  prac- 
tically the  same,  if  not  identical,  whether  the  salts  are  laid  down 
under  normal  conditions  (ossification)  or  under  pathological  condi- 
tions. With  the  blood  continually  passing  between  the  bones  and 
the  calcified  areas,  the  composition  of  the  two  must  inevitably  become 
similar  or  identical.  This  may  be  shown  bj'  a  table  giving  the  pro- 
portion of  inorganic  salts  found  by  analysis  of  normal  bone,  and  the 
proportion  found  in  calcified  materials.^ 


Pathological  Calcification. 
Bovine  tuberculosis 

'•  "  (softened  gland)    . 

Human  tuberculosis 

Calcified  nodule  in  thyroid 
Thrombus,  human 

Nokmal  Ossification. 
Human  bone   (Zalesky)        .... 

"      (Carnot") 

"      (Carnot)    

Ox  bone   (Zalesky) 

"       "       (Carnot)      


Mg3(P04)2. 


0.84 

0.9 

1.2 

1.5 

1.2 

0.85 

1.1 


1.04 
1.57 
1.75 
1.02 
1.53 


CaCOa. 


12.8 
13.1 
11.7 
7.6 
10.1 
13.4 
11.9 


±12.8 

10.1 

9.2 

il.9 


CaaCPO*),. 


85.9 
85.4 
86.4 
90.6 
87.8 
85.4 
86.5 


83.8 
87.4 
87.8 
86.1 
85.7 


Iron  may  be  present  in  pathological  calcification,  and,  according  to 
Gierke,*  in  the  fetus  the  entire  skeleton  contains  iron  as  far  as  it  has 
calcified,  most  at  the  points  of  active  ossification.  This  statement 
has  been  questioned  by  Hiick  and  others,  who  believe  that  most  of 

2a  MacCordick  (Lancet,  Oct.  18,  1913)  has  advanced  the  interestinij  hypothesis 
tliat  calcific  deposits  during  life  exist  mostly  as  soft  masses,  like  unset  mortar. 
Only  when  sufficient  accumulation  of  CO,  occurs,  as  after  death,  or  in  tlie  center 
of  large  areas  of  low  vitality,  such  as  fibroids,  do  the  deposits  become  hardened: 
e.  g.,  in  a  gangrenous  leg  the  calcified  vessels  arc  stilT  and  brittle,  wliile  higlier 
up  in  tlie  living  tissues  they  are  soft  and  ])liable.  This  would  explain  why  we  do 
not  more  often  observe  fractures  of  calcified  arteries.  As  yet  this  hy])o(hesis  has 
not  received  the  critical  tests  its  imi)ortance  deserves.  If  true  it  will  explain 
the  cases  of  extensive  calcification  of  the  pericardium  in  which  the  heart  is  so 
encased  that  function  would  seem  impossible  if  the  deposit  were  rigid  during 
life.  (See  Trans.  Chicago.  Pathol.  Society,  1911  (8),  109,  for  consideration  of 
pericardial  calcification.)  However,  Klot/  (.lour.  Med.  Res.,  191()  (34),  495) 
lias  (|U(!stioned  the  correctness  of  MacCordick's  views  on  the  basis  of  the  occa- 
sional occurrciice  of  fractures  of  calcified  arteries,  but  without  experimental  evi- 
dence contradicting  MacCordick. 

3  Wells,  lor.  fit. 

4  Vircliow's  Arch.,  1902   (107),  318.       • 


CALCIFICATION  437 

the  iron  demonstrable  in  noimal  ossification  is  the  result  of  an  arti- 
fact, for  calcium  deposits  seem  to  have  a  great  affinity  for  iron.  Be- 
cause of  this,  pathological  calcium  deposits  take  up  iron  from  old 
hemorrhages  in  the  vicinity,  and  so  in  man}-  areas  where  there  have 
been  hemorrhages,  especially  in  the  vicinity  of  elastic  tissue,  there 
occur  actual  ''calcium-iron"  incrustations.'^  S.  Ehrlich"  states  that 
elastic  fibers  in  the  vicinity  of  hemorrhages  take  up  the  iron-contain- 
ing derivative  of  tlic  blood-pigment,  and  this  acts  as  a  mordant  for 
subsequent  calcium  deposition.  The  presence  of  iron  in  normal  ossi- 
fication is  supported  by  Sumita'^  and  Eliasscheff.*  In  the  so-called 
iron-lime  lung  Gigon  ^  found  l>ut  a  trace  of  calcium  and  much  sodium 
and  potassium. 

Structure  of  Calcified  Areas. — As  before  mentioned,  in  calcifi- 
cation there  is  not  the  same  uniform  infiltration  of  the  ground  sub- 
stance with  lime  salts  that  occurs  in  bone,  yet  the  calcified  area  is  pos- 
sessed of  a  ground  substance  of  organic  material  which  does  not  dis- 
solve in  acids  that  remove  the  salts.  There  is  no  definite  ratio  be- 
tween the  lime  salts  and  this  albuminoid  matrix,  however.  At  first 
the  salts  occur  in  granules,  which  may  become  fused  to  a  greater  or 
less  degree.  It  has  been  thought  b}'  some  that  the  deposition  occurs 
in  the  form  of  ' ^ calcospherites." 

These  are  small  calcareous  bodies,  iisiially  of  concentric  structure,  which  were 
first  described  by  Harting.  Tliey  appear  to  occur  widely  distributed  in  normal 
tissues,  both  animal  and  plant,  and  seem  to  be  the  result  of  the  formation  of 
insoluble  calcium  salts  in  the  presence  of  some  organic  substances,  just  as  urinary 
and  other  concretions  are  formed  about  an  organic  nucleus.  If  calcium  chloride 
and  soluble  carbonates  are  allowed  to  combine  very  slowly  to  form  calcium  car- 
bonate in  a  solution  of  e£rg-albumen,  these  or  indistinguishable  bodies  are  formed, 
which  on  being  dissolved  are  found  to  possess  an  organic  stroma  that  exhibits  a 
marked  affinity  for  any  pigmentary  substance  that  may  be  present.  Apparently, 
Avhen  the  proper  concentration  exists,  the  salts  in  crystallizing  hold  between  the 
crystals  tlie  albuminous  substances  by  which  they  are  surrounded.  Dastre  and 
Morat  believe  that  the  substratum  is  lecithin,  which  others  have  found  occupying  a 
similar  place  in  prostatic  concretions.  Calcospherites  have  been  found  in  tumors, 
in  cystic  cavities,  and  in  bodies  with  beginning  decomposition.  It  may  be  men- 
tioned in  passing  that  Littlejohn  »  observed  the  abundant  formation  of  calcium 
l)hosphate  crystals  in  bodies  that  had  been  immersed  for  some  time  in  sea  water. 
Oliver  has  found  calcospherites  in  the  tissues  of  a  cancer  of  tlie  breast.  Pettit  lo 
found  calcospherites  in  a.  sarcoma  of  the  maxilla,  presenting  insensible  transi- 
tions into  the  substance  of  the  osseous  tissue,  and  he  suggests  the  possibility  that 
the  calcospherite  formation  may  be  related  to  the  formation  of  bone.  It  seems, 
however,  that  tliey  are  probably  more  closely  related  to  the  formation  of  the 
shells  of  invertebrates,  which  are  largely  composed  of  carbonates  in  crystalline 
structure  with  an  organic  ground  substance  between  them,  and  very  little  phos- 
phate indeed. 

sSee  Gigon,   Ziegler's   Beitr.,   1912    (55),   4G;    Sprunt,  Jour.   Exp.   Med.,    1011 
(14),  50. 
«  Cent.  f.  Pathol..  1000    (17),  177. 
■Virchow's  Arch.,  1010    (200).  220. 
sZiegler's  Beitr.,   1011    (50),   14.3. 

9  Edinburgh  ^Nled.  Jour.,  100.3    (13),  127. 

10  Arch.  d.  Anat.  Micros.,  1897    (1),  107. 


438  dALCIFICATION,    CONCRETIONS,    AND    INCRUSTATIONS 

OCCURRENCE  OF  PATHOLOGICAL  CALCIFICATION 

As  far  as  we  know,  calcification  seldom  occurs  in  normal  tissue, 
except  in  the  formation  of  bone.  Often  the  infiltrated  tissue  is  com- 
pletely dead,  as  in  infarcts,  organic  foreign  bodies,  caseous  areas,  and 
particularly  in  old  inspissated  collections  of  pus.  It  may  be  said 
that  any  area  of  dead  tissue  that  is  not  infected,  and  that  is  so  large 
or  so  situated  that  it  cannot  be  absorbed,  will  probably  become  infil- 
trated with  lime  salts.  i\Iost  frequently  calcified,  next  to  totally  ne- 
crotic tissues,  are  masses  of  scar-tissue  that  have  become  hyaline  sub- 
sequent to  the  shutting  off  of  circulation  in  the  scar  by  contraction 
of  the  tissue  about  the  vessels.  Elastic  tissue  also  seems  prone  to  an 
early  calcification,  and  it  is  not  uncommon  to  see  the  elastic  laminaB 
of  small  arteries  calcified  in  an  apparently  selective  manner.  A  pe- 
culiar form  of  calcification  is  that  frequently  found  in  ganglion-cells 
of  the  brain  which  have  become  degenerated  or  necrotic,  particularly 
in  the  vicinity  of  old  hemorrhages;  the  cells  become  infiltrated  with 
lime  salts  until  a  complete  cast  of  the  cell,  with  dendrites  and  axis- 
cylinder  well  impregnated,  is  formed.  The  calcification  of  renal  epi- 
thelium obtained  experimentally  by  temporary  ligation  of  the  renal 
vessels  or  by  the  administration  of  certain  poisons,  is  more  closely 
related  to  the  formation  of  ordinary  urinary  eoneretions  than  to  tissue 
calcification,  the  calcium  being  present  as  the  phosphate  only.^^  Cal- 
cification of  epithelial  cells  does  occur,  however,  and  seems  to  be  pre- 
ceded by  hyaline  changes,  in  which  hyaline  substance  the  calcium  is 
later  deposited,  as  in  epithelial  pearls,  for  example. 

Metastatic  Calcification. — What  is  perhaps  the  only  exception  to 
the  rule  that  some  form  of  tissue  degeneration  is  required  before  cal- 
cification occurs,  is  the  ''metastatic  calcification"  of  Virchow.^-  In 
conditions  with  much  destruction  of  bone,  as  osteomalacia,  caries, 
osteosarcoma,  etc.,  deposits  of  lime  salts  have  been  found  distributed 
diffuseh^  in  various  organs,  particularly  in  the  lungs  and  stomach. 
As  much  as  13.38  per  cent,  of  the  dry  weight  of  the  lung  and  12.15 
per  cent,  of  the  kidney  have  been  found  as  CaO  in  such  a  case.^--'' 
As  there  is  no  evidence  that  these  organs  have  been  the  site  of  any  dif- 
fuse tissue  necrobiosis  before  the  calcification  occurred,  it  seems  prob- 
able that  the  deposits  have  been  made  in  practically  or  quite  normal 
organs,  because  of  oversaturation  of  the  tissue  fluids  by  calcium  salts. 
The  fact  that  the  lung  and  stomach,  and  also  to  a  less  degree  the  kid- 
ney, are  picked  out,  suggests  that  the  calcification  is  related  to  the 
fact  that  in  these  same  organs  we  have  the  excretion  of  acids  into 
their  cavities,  which  leaves  the  fluids  in  the  substance  of  the  organs 

11  Jour.  Med.  Researeli.  1011    (2.")),  :17.1. 

12  Virohow'a  Arrli..  IS.'i.'S  (S),  10.3:  review  by  Koekel.  Dent.  .\reh.  klin.  IMed., 
1899  (64),  332.  Biblioj,r,aphv  and  review  bv  Wells,  Arch.  Int.  Med.,  1015  (15), 
574. 

i-'a  Virchow's  .\rcli.,  1000    (107),  112. 


,,,;.;  U/NVA'V  O/'  77//;   /'A'OCESS  OF  CALCIFIC.iTIOy  439 

T  1  .,lk.,lino  and  an  increase  in  the  alkalinity  of  the 
oorresponchngly  ^^^'^.^"\''  'Vt'^ecidedlv  less  soluble.  In  the  stomach 
fluids  makes  the  calcumi  ^f  ^^J^^^^'^  ^^^ter-landular  tissue  about 
the  ealeium  deposits  ^^  ^J^i::^:':..^  corresponding 
the  upper  portion  of  the  ?U>^^*^  .^.^^osed  to  secrete  the  acid.  Pre- 
to  the  parietal  cells  which  '^V^     "^^^j  ^''' J,,,,,t   of   calcium   in   the 

blood  IS  too  slignt  to  ue  uiiu  skeleton,  precipita- 

attemrted  to  include  'l;^"^!"'*',!^^'^.^^  ^cX  ,  h  oVi.in  of  the 
in  old  age  in  the  metastatic  =«1"«°'^*'°°'' *'";""  bably  dependent 
salts  to  the  senile  f -'T'*-"  .^^ ^''™  :;„tltfon  oft^^^^^  tis- 

:^:JZ:^  rtrs:r iSe\ot:"Tchan,e  ....  .een.  to  he 

rCo'lra^tpoSnffaeto't^tt:  solTtion  of  ealeinn.  salt.  ,n 

111  rt  the  Uo  d%s  tcreS  hviniectin,  or  feedin.  calcium 
:  t     deposi«ons  ot  calcium  salts  may  take  place  m  injured  tissues, 
or  even  in  normal  tissues,  as  in  Tanaka's  experiments.  • 

CHEMISTRY  OF  THE  PROCESS  OF  CALCIFICATION 
Tn  analv^iuK  the  etiological  factors  in  the  production  of  pathologi- 

bo  at  an  phosphate  themselves,  or  as  ealcium-ion-protem  com- 
^o  nds  or  perhaps  both.  This  suspension  or  solution  is  an  unstable 
eondt;.,  poss  ble  only  because  of  the  extremely  small  proportion  o 
calctm"n  ^e  blood  (about  1  :in,O0O),  and.  therefore,  capable  of 
be  n"mcrtl  rown  bv  increased  alkalinity  of  the  blood,  changes  m  the 
prtSiiis  or  CO,  content,  or  changes  in  the  quantity  or  composition 

r  u     1  •  „„+  "  -NT    T5    «5p>innAt  has  described  a  case  with 

,eS^/^;ST„„°l,':;,'fcZ    no.  ^L  »^^  „™..K-  allele,  in 

physiol.  Chem'.,  1913    (S5).  324.  iqnTn)     1 

"^:STzSi^^r\^^r^rnif:^':  i«  a.Vo  Kata.,  Be...  pa.„. 
Anat.,  1914   (57),  516. 


440  CALCIFICATION,    CONCRETIOXS,    AND    IXCRUSTATI0X8 

of  tlie  ealciiim  salts.  It  is  probable,  from  the  work  of  Barille,  that 
the  calcium  of  the  blood  exists  as  a  soluble  complex  double  salt,  tri- 
basic  calcium-earbon-phosphate  (PoOgCa,!!, :  2COo(C03H)2Ca),  this 
compound  being  possible  because  of  an  excess  of  CO,. 

(2)  Retrog-ressive  changes  in  the  tissues  are  a  sine  qua  non.  Hya- 
line degeneration,  the  chemical  nature  of  which  is  not  understood,  is 
a  veiy  favorable  condition,  as  also  is  necrosis  when  absorption  is 
deficient. 

(3)  In  the  areas  that  are  to  become  calcified  the  circulation  is 
very  feeble,  the  blood  plasma  seeping  through  the  tissue  as  through 
any  dead  foreign  substance  of  similar  structure,  without  the  presence 
of  red  corpuscles  to  permit  of  oxidative  changes. 

We  may,  therefore,  imagine  that  the  dei)osition  of  calcium  salts 
in  such  areas  of  tissue  degeneration  depends  upon  one  or  more  of  the 
following  conditions : 

(1)  Increased  alkalinity  or  decreased  CO,  in  the  degenerating 
tissues,  causing  precipitation  of  the  inorganic  salts  in  the  fluids  seep- 
ing slowly  through  them. 

(2)  Utilization  of  the  protein  of  the  fluids  by  the  starved  tissues 
so  completely,  because  of  its  slow  passage  through  them,  that  the 
calcium  cannot  be  held  longer  in  solution. 

(3)  The  formation  within  tlie  degenerated  area  of  a  substance  or 
substances  having  a  special  affinity  for  calcium. 

(4)  Production  of  a  physical  condition  favoring  the  local  absorp- 
tion of  salts,  the  least  soluble  salts  accumulating  in  excess. 

The  first  of  these  conditions  seems  to  come  into  play  especially  in 
metastatic  calcification,  already  discussed.  We  have  no  evidence  that 
in  degenerating  tissues,  much  less  in  normal  ossification,  there  is  an 
alkaline  reaction  developed ;  but  rather  the  contrary^  an  acid  reaction 
is  more  usual.  But,  as  explained  below,  decrease  in  the  CO,  content 
in  calcifying  tissues,  especially  when  combined  with  other  changes, 
may  be  of  importance. 

Lichtwitz  ^^  especially  has  laid  emphasis  on  the  possible  part  played 
by  changes  in  the  proteins  in  inducing  calcification.  He  advances 
the  idea  that  precipitation  of  the  colloids  in  the  degenerated  area, 
as  in  caseation,  decreases  the  amount  of  crystalloids  which  can  be 
held  in  solution,  wherefore  the  least  soluble  salts,  tliose  of  calcium, 
are  precipitated;  by  laws  of  osmotic  pressure  more  calcium  in  solu- 
tion will  then  enter  to  establisli  ('()uilibrium,  be  precipitated,  and 
make  way  for  more  calcium,  until  the  amount  of  deposit  prevents 
further  osmotic  diffusion.  Altliough  suggestive  in  regard  to  patho- 
logical calcification,  and  probably  of  importance  in  the  fonnation 
of  concretions,  this  conception  is  difficult  to  aiii)ly  to  normal  ossifi- 
cation ;  also  in  pathological  calcification  one  would  exjiect  precipi- 
tation of  calcium  to  occur  in  tlie  outermost  surface  of  the  degener- 

11  Doiit.  mcd.  Wocli.,  1010    (aO),  704. 


l'()l!M\n<)\   OF  CM.illU   SOM'S  441 

ated  area,  soon  loading  to  a  sliell  of  inorganic  material  which  would 
limit  the  deposition. 

The  possibility  of  the  formation  of  calcium-binding  substances 
within  the  degenerated  area  has  always  seemed  the  most  attractive, 
and  has  received  the  most  attention  by  investigators.  Of  the  special 
substances  that  might  be  present  in  such  areas  that  would  have  a 
high  afifinity  for  calcium,  phosphoric  acid  usually  receives  first  con- 
sideration, since  it  is  as  phos])hate  tliat  most  of  the  calcium  is  bound, 
and  also  since  the  possible  sources  of  phosphoric  acid  in  decomposed 
nucleoproteins  and  lecithin  are  so  obvious.  Less  considered  in  the 
past,  fatty  acids  offer  another  possibility,  especially  in  view  of  the 
fatty  degeneration  that  so  frequently  precedes  calcification.  Proteins 
might  also  be  formed  that  Avould  combine  calcium,  especially  deutero- 
albumose,  which  Croftan  ^^  states  has  a  high  degree  of  affinity  for 
calcium,  and  which  would  be  present  in  areas  undergoing  autoly- 
sis. 

Formation  of  Calcium  Soaps. — In  favor  of  the  possibility  that 
the  calcium  is  first  bound  as  soaps  are  the  following  facts :  Calcifica- 
tion occurs  chiefly  in  places  where  fatty  degeneration  has  occurred, 
such  as  tubercles,  atheromatous  vessels,  etc.  In  fat  necrosis  fatty 
acids  are  formed,  which  soon  combine  with  calcium  to  form  calcium 
soaps.  Virchow  observed  calcification  in  the  form  of  soaps  in  a 
lipoma,  and  Jaeckle  ^*  found  that  a  calcifying  lipoma  contained  29.5 
per  cent,  of  its  calcium  in  the  form  of  calcium  soaps.  Klotz  ^^  ob- 
tained staining  reactions  in  calcifying  tissues  that  suggested  the  pres- 
ence of  soaps,  which  he  also  extracted  by  solvents,  and  he  strongly 
urges,  as  the  first  step  in  the  formation  of  pathological  calcified 
masses,  that  the  calcium  is  first  laid  down  as  soaps,  afterward  under- 
going a  transformation  into  the  less  soluble  phosphate  and  carbonate. 
Fischler  and  Gross  -"  also  obtained  microchemical  reactions  for  soaps 
in  the  margins  of  infarcts  and  in  atheromatous  areas,  but  not  in 
caseous  areas;  they  therefore  consider  that  calcium-soap  formation  is 
an  important  step  in  the  process  of  pathological  calcification,  but 
that  it  is  not  essential.  The  value  and  the  interpretation  of  the  his- 
tological evidence  of  the  participation  of  calcium  soaps  is,  however, 
open  to  question. 

On  the  other  hand,  Wells,-^  studying  large  quantities  of  material 
chemically,  found  at  most  doubtful  traces  of  calcium  soaps  in  calci- 
fying matter,  even  in  the  earliest  stages,  and  also  very  small  amounts 
of  other  soaps  or  fatty  acids,  and,  therefore,  questions  the  occurrence 
of  calcium  soaps  as  an  essential  step  in  calcification,  although  not 
doubting  that  under  certain  conditions  (e.  g.,  calcifying  lipomas,  fat 

I'.Tour.  of  Tuberculosis.  in03   (5),  22. 
isZeit.  physiol.  Chem.,  inn2    (36).  .53. 

19  Jour.  Exper.  :Nred.,  ion.5    (7),  633;    1906    (S),  322. 

20  Ziegler's  Beitr.,  1005    (7th  suppl.).  339. 

21  See  review  in  Arch.  Int.  Med.,  1911    (7),  721. 


442  CALCIFIC ATlOy,    COyCREllOyfi,    A.VZ)    lyCRU STATIONS 

necrosis)  this  may  occur.  In  calcification  at  all  stages  the  propor- 
tion of  calcium  carbonate  and  phosphate  was  found  quite  constant, 
and  exactly  the  same  as  in  normal  bone;  namely,  in  the  proportion 
expressed  by  the  formula  3(Ca.5(POJo)  :  CaCO,,  which  Hoppe-JSeyler 
advanced  to  express  the  composition  of  the  salts  of  bone.  Hence  it 
seems  probable  that  there  are  no  essential  differences  between  the 
processes  of  ossification  and  pathological  calcification,-^''  and  there 
seems  to  be  as  yet  no  reason  for  assuming  that  in  the  former  calcium 
soaps  constitute  an  essential  step  in  the  process. 

Phosphoric  Acid  in  Calcification. — It  has  generally  been  assumed 
that  in  normal  ossification  the  calcium  is  combined  by  phosphoric 
acid,  which  probably  is  derived  from  the  cartilage  cells,  possibly 
through  autolysis  of  the  nucleoproteins  or  some  similar  process.-- 
Grandis  and  Mainini,-^  by  using  mierochemical  methods,  thought  that 
thej^  found  evidence  that  the  phosphorus  of  ossifying  cartilage  is 
converted  from  an  organic  combination  into  an  inorganic  form 
(PoOJ,  which  then  takes  up  calcium  from  the  blood.  The  methods 
used  have  been  questioned,  and  Pacchioni,-*  from  his  studies,  was  in- 
clined to  the  opinion  that  the  calcium  entered  the  cartilage  already 
combined  as  phosphate.  Wells  implanted  into  the  abdominal  cavity  of 
rabbits  various  tissues  that  had  been  killed  and  sterilized  by  boil- 
ing, and  found  that  tissues  rich  in  nucleoproteins  showed  no  tendency 
to  take  up  calcium  in  greater  amounts  than  did  tissues  poor  in  nucleo- 
proteins, which  result  speaks  against  the  idea  that  phosphoric  acid 
derived  from  nucleic  acid  combines  the  calcium.  On  the  other  hand, 
implanted  dead  cartilage  soon  became  thoroughly  impregimted  with 
calcium  salts,  which  seemed  to  be  deposited  in  the  same  proportion 
as  to  carbonate  and  phosphate  as  in  bone. 

Physical  Absorption  of  Calcium  Salts. — As  there  could  be  no 
question  of  "vital  activity"  on  the  part  of  this  boiled  cartilage,  it 
seems  most  probable  that  there  exists  in  cartilage  a  specific  absoi"p- 
tion  affinity  for  calcium  salts,  similar  to  the  absorption  affinity  that 
ITof meister -^  observed  exhibited  by  other  organic  colloids  (gelatin 
disks)  toward  various  crystalline  substances  in  solution.  It  is  of  sig- 
nificance that  the  substances  in  which  calcium  is  deposited  are,  in 
most  instances,  of  similar  character,  being  homogeneous  and  often 
hyaline,  although  of  the  most  varied  chemical  composition;  in  other 
words,  they  agree  much  more  in  physical  tlian  in  chonical  stnicture. 
Also  we  find  tliat  liyalino  tissues  witli  an  affinity  for  calcium  often 
exhibit  a  similar  affinity  for  othei-  substances,  such  as  pigment  and 

'.iiii  Dyps  tliat  stain  the  bones  \\\wn  fed  to  livinji:  animals  (madder)  also  stain 
patholof^ical  calcific  deposits    (Macklin,  Anal,   rvccord.   1!)17    (11).  387). 

22  TTanes,  who  observed  that  the  phosphatids  disappear  from  tlie  liver  of  the 
developing  chick,  supfjests  this  as  a  source  of  the  iijiosphoric  acid  rcijuired  for 
ossification   (Jour.  Exper.  ]\led.,  1012   (10),  512). 

23  Arch,  per  la  sci.  ]\Ted.  Torino,  1000    (24),  07. 
2'»  Jahrb.  f.  Kinderheilk.,  1002   (56).  327. 

2.'.  Arch,  exper.  Path.  n.  Pharm.,  1801    (28),  210 


OSTEOMALACIA  443 

iron.-"  Ilofmeister  advances  the  hypothesis  that  when  the  cartilage 
or  other  matrix  becomes  saturated  with  calcium  salts,  any  decrease 
in  COo  content  of  the  solution  will  lead  to  a  precipitation  of  calcium 
salts,  thus  restoriiijj;  to  the  cartilage  its  power  of  absorbing  more 
calcium  salts  whenever  the  fluid  comes  to  it  with  a  higher  degree  of 
saturation  with  calcium  salts  and  COg.  This  hypothesis  is  in  har- 
mony with  Barille's  observation  that  when  the  CO2  is  reduced  the 
complex  car])oii-pliosphate  of  calcium  precipitates  a  mixture  of  car- 
bonate and  phosphate  in  the  same  proportions  as  found  in  bones  and 
calcific  deposits  generality.  The  fact  that  this  ratio  (10  to  15  per 
cent.  CaCO.j  and  85  to  90  per  cent.  Ca;j(P04)o),  is  found  in  all  stages 
of  calcification,  is  entirely  in  favor  of  the  above  hypothesis,  and 
opposed  to  the  idea  that  any  special  chemical  precipitant  formed  in 
the  calcifying  area  is  responsible  for  the  deposition  of  calcium. 
Taken  all  in  all,  the  evidence  seems  in  favor  of  the  view  that  normal 
ossification  and  pathological  calcification  (except  metastatic  calcifica- 
tion and  the  calcification  of  fat  necrosis  and  other  areas  of  necrotic  fat 
tissue)  depend  more  upon  physico-chemical  factors  and  variations  in 
CO2  concentration  than  upon  the  presence  of  chemical  precipitants 
in  the  tissues. 

OSTEOMALACIA  27 

In  this  condition  the  quantity  of  inorganic  salts  in  the  bone  is 
greatly  decreased,  while,  at  the  same  time,  their  place  is  taken  in 
part  by  new-formed  osteoid  tissue ;  as  a  result,  the  proportion  of  the 
weight  of  the  bone  formed  by  inorganic  salts  is  reduced  to  as  low  as 
20  to  40  per  cent.,  instead  of  being  from  56  to  60  per  cent.,  as  in  nor- 
mal bone.  This  has  suggested  that  the  cause  of  the  disease  may  be 
a  solution  of  the  lime  salts  by  some  acid,  but  Levy  -^  found  that 
in  osteomalacia  the  proportion  of  calcium  carbonate  and  phosphate 
in  the  bones  remains  constant,  as  also  does  the  proportion  of  cal- 
cium and  phosphoric  acid ;  if  the  decalcification  occurred  through 
solution  by  lactic  or  other  acids,  the  carbonate  should  be  decora- 
posed  first,  whereas  the  lime  salts  seem  to  be  taken  out  as  mole- 
cules of  calcium  carbonate-phosphate ;  i.  e.,  in  the  same  propor- 
tion as  they  exist  in  the  bone.  On  the  other  hand,  it  has  been  found 
in  Pawlow's  laboratory  that  dogs  kept  for  long  periods  after  a  pan- 
creatic fistula  has  been  established,  develop  a  condition  resembling 
osteomalacia, "°  which  would  seem  most  reasonably  explained  as  due 
to  the  constant  loss  of  alkali  in  the  pancreatic  juice.  Furthermore, 
investigation  of  Levy's  objection  to  the  acid  solution  theory  has  led 

26  See  Sprunt,  Joiir.  Exp.  Med.,  1011    (14),  i^O. 

27  See  also  review  in  Albn  and  Neuberp's  "^rineralstofTwechsel,"  Berlin,  lfl06, 
pp.  124-127;  biblioirraphy  by  Zesas,  Cent.  Crenz.  :\led.  u.  C'hir.,  1007  (10),  801; 
full  discussion  bv  ]\IcCru'dde'n,  Arch.  Int.  INIed.,  1010  (•>),  riW:   1012   (0),  27.3. 

28Zeit..  physiol.  Chem.,  1804    (10),  2.30. 

29  Babkin,  Zeit.  Stoffwechsel,  1010  (11),  .561;  Looser,  Verh.  Dent.  Path.  Gesell., 
1007    (11),  201. 


444 


CALCIFICATION,    COXCRETIOXS,    AXD    INCRUSTATIONS 


to  the  observation  that  when  mixtures  of  calcium  carbonate  and  phos- 
phate are  in  colloids  they  are  dissolved  at  equal  rates.-''^  McCrudden 
found  tliat  in  the  bones  in  human  osteomalacia,  together  with  the  de- 
crease in  calcium  there  is  an  increase  in  the  magnesium  ^°  and  sul- 
phur, because  of  newly  deposited  tissue  poor  in  calcium.  Histologic- 
ally, absorption  seems  to  depend  largely  upon  a  direct  eating  out  of 
bone  tissue,  both  organic  and  inorganic  substance,  by  osteoclasts 
(Cohnheim),  followed  by  a  formation  of  an  uncalcified  osteoid  tis- 
sue. (Senile  osteoporosis  differs  chiefly  in  that  no  new  osteoid  tissue 
is  formed.)  According  to  Dibbelt  ^^  when  osteomalacia  is  experimen- 
tally induced  in  pregnant  dogs  and  then  recovery  is  allowed  to  take 
place,  the  decalcified  bone  substance  present  in  the  active  stage  does 
not  become  calcified,  but  is  absorbed  and  replaced  by  new  bone. 

Studies  of  metabolism  in  osteomalacia  have  shown  a  loss  of  calcium 
by  the  body,  especially  in  the  urine,  as  shown  by  the  following  table 
given  by  Goldthw^ait  et  al. :  ^' 


Limbeck 

Neumann 

Goldthwait 

CaO  in  urine   (gm.) 

CaO  in  feces 

1.773 
3.S34 

3.859 
1.800 

Total  excreted 

Total  in  food 

5.607 
2.965 

2.965 

11.65 
11.26 

5.66 
4.56 

Loss  of  CaO 

0.39 

1.10 

McCrudden  also  found  a  considerable  retention  of  nitrogen  and 
sulphur,  which  may  be  retained  in  the  new-formed  osteoid  tissue ; 
magnesium  is  also  retained,  probably  being  substituted  for  calcium 
in  the  bones.  It  is  known  that  when  magnesium  and  strontium  are 
given  to  growing  animals  they  will  partially  replace  the  calcium  in 
the  bones,^-''  while  it  is  said  by  Etienne  ^^  that  excessive  feeding  of  cal- 
cium itself  leads  in  time  to  decalcification  of  the  bones.  Zuntz  ^* 
found  the  respiratoiy  metabolism  in  osteomalacia,  within  normal  lim- 
its, but  tending  to  be  low;  protein  metabolism  shows  nothing  strik- 
ing, but  there  is  a  high  excretion  of  phosphoric  acid  througli  the  feces. 

Castration  of  women  with  osteomalacia  has  been  frequently,  but 
not  always,  followed  by  improvement  or  recovery,^^"^  and  Neumann, 
and  also  Coldthwait,  have  found  that  in  these  cases  the  calcium  loss  is 
rejjlaced  by  a  marked  calcium  retention  after  the  0])eration.  AVhat 
tlie  relation  of  the  ovaries  to  calcium   metabolism  or  to  osteomalacia 

2naKran/.  and  Lioso<,'anfr,  Dent.  Monat.  Zalniheilk.,  1914,  p.  62S. 

30  Corrolmratod  by  C'ai)pez/.U(>li,  Biocliem.  Zeit.,   1909    (16).  355. 

31  Arbeit.  Patli.  Tnst.  Tuhinf,'en,  1911    (7),  559. 

32Goldlli\vait,    Painter,    Osfjood    and    I\IcCr\idden,    Amcr.    .lour.    I'livsiol..    1905 
(14),  389. 
32a  See  Lelinerdt,  Zeit.  exp.  Med.,   1<)1;5    (1).   175. 
33  Jour.   I'livsiol.  et  Path.,   1912    (14).   lOS. 
34Areli.   f.  fiyn.,   1913    (99),    145. 
34a  Bibliography  by  Schnell,  Zeit.  Oeb.  u.  (Jyn.,  1913    (75),  178. 


h'lChlJT.S 


445 


may  be  has  not  yet  been  ascertained,  ^charf e  ^■'  and  Bucura  ^"  both 
state  that  tliere  are  no  cliaracteristic  or  constant  structural  altera- 
tions in  the  ovaries  in  osteonuUacia.  McCrudden  "'  found  that  the 
improvement  in  calcium  metabolism  observed  after  castration  may 
be  but  temporary,  and  therefore  believes  that  the  primary  cause  of 
the  disease  does  not  lie  in  the  ovaries.  He  is  of  the  opinion  that  re- 
peated drains  on  the  calcium  of  the  bones,  incited  most  often  by 
pregnancy,  occasionally  by  tumors,  sometimes  by  unknown  causes, 
result  in  an  excessive  reaction  to  the  stinuili,  so  that  eventually  the 
losses  become  too  great  to  be  made  up ;  that  is,  osteomalacia  is  an 
exaggei-ation  of  a  normal  process  resulting  either  from  excessive 
stimulation  of  that  process,  or  a  failure  to  recover  when  the  stimulus 
ceases.  The  beneficial  effects  of  castration  are  probalily  ascribable 
chiefly  or  solely  to  the  prevention  of  pregnancy.  Osteitis  deformans 
seemed  to  be  a  localized  osteomalacia.  The  relation  of  the  adrenals  to 
osteomalacia  advocated  by  Bossi,^*  is  of  questionable  significance,  and 
there  is  no  definite  evidence  as  to  any  relation  of  exophthalmic  goiter  ^^ 
or  the  parathyroids,^"  although  hyperplasia  of  the  parathyroids  has 
been  described." 

RICKETS  i-^ 

As  with  osteomalacia,  chemical  studies  of  the  bones  in  rickets  have 
thrown  little  light  upon  the  etiology  or  pathogenesis  of  this  condition. 
As  the  following  table  (taken  from  Vierordt*^)  shows,  there  is  a 
marked  deficiency  in  the  proportion  of  inorganic  salts  in  the  bones  in 
rickets.  The  proportion  of  the  different  salts  seems  to  be  quite  the 
same  as  in  normal  bone. 


Normal  bone 

of  a  two 

months  old 

child 

Rachitic  bones 

Tibia. 

Ulna. 

Femur. 

Tibia. 

Humerus. 

18.88 
81.12 

Ribs. 

37.19 
62.91 

Vertebras. 

Inorganic    matter 

Organic     substance  .... 

65.32 
34.68 

57.54 
1.03 
6.02 
0.73 

33.86 
0.82 

64.07 
35.93 

56.35 
1.00 
6.07 
1.65 

34.92 
1.01 

20.60 
79.40 

14.78 
0.80 
3.00 
1.02 

72.20 
7.20 

33.64 
66.36 

26.94  \ 

0.81   / 

4.88 

1.08 
60.14  \ 

6.22  / 

32.29 
67.71 

Calcium  phosphate    .... 
Magnesium    phosphate.  . 
Calcium   carbonate    .... 

Soluble    salts     

Collagen   (or  ossein) .  .  . 

15.60 
2.66 
0.62 

81.22 

445. 


35  Cent.  f.  Gyn.,  1900   (24),  1216. 

36Zeit.  f.  Heilk.,  1907   (28),  209. 

37Amer.  Jour,  of  Physiol.,  1906    (17),  211. 

38Zent.  f.  Gyn.,  1907   (31),  69  and  172. 

39Tolot  and  Sarvonat,  Rev.  d.  Med.,  1906    (26 

4"Erdheim,  Cent.  nied.  Wiss.,  1908    (46),   163. 

■41  Bauer.  Frankfurter  Zeit.  Pathol.,  1911    (7),  231. 

•12  Complete  literature  and  full  discussion  bv  Pfatmdler.  -lalir.   f. 
1904    (60),  123:  also  see  Albu  and  Xeuberg.  ".Mineralstotl'weclisel," 
pp.   119-124;    symposium   in  the  Verhandl.   Deut.   Path.  Gesellseh., 
Metabolism  studies  by  Meyer,  Jahrb.  Kinderheilk.,  1913   (77),  28. 

43  Xothnagel's  System,  vol.  7,  part  ii,  p.  21. 


Kin 

Ber 

1909 


dcrheilk.. 
n.  1906, 
(13),   1. 


446  CALCIFICAriOX,    COyCKETIOXH,    AXD    IXCRL  STATIONS 

INIore  modern  analyses**  show  a  relative  increase  in  water  and 
magnesium,  with  a  persistence  of  the  normal  ratio  of  calcium  phos- 
phate and  carbonate.**''  Cattaneo  *"^  finds  the  increase  in  magnesium 
to  var}-  in  different  parts  of  the  skeleton,  being  greatest  in  the  ribs. 
The  blood  of  children  with  rickets  shows  greater  variations  from  the 
usual  CaO  content  (8-10  mg.  per  100  c.c.)  than  are  found  in  normal 
children  ( Asehenheim)  .*'"' 

As  an  essential  difference  from  osteomalacia  is  the  fact  that  in 
rickets  there  is  a  failure  on  the  part  of  the  osteoid  tissues  to  calcify, 
Avhereas  in  osteomalacia  absorption  of  calcified  tissue  takes  place 
with  subsequent  substitution  by  osteoid  tissue.  Furthermore,  in 
rickets  the  deficiency  in  calcium  is  said  to  be  present  only  in  the 
bones/'^  whereas  in  osteomalacia  the  soft  tissues  are  also  poor  in  lime 
salts.  According  to  Schmorl  *'^  the  first  structural  abnonnality  in 
rickets  is  a  failure  to  lay  on  calcium  by  small  islands  of  cartilage  in 
the  zone  of  preparatory  calcification. 

None  of  the  various  hypotheses  as  yet  advanced  to  explain  this 
defective  ossification  has  satisfactorily  explained  all  the  observed 
facts.  That  a  deficiency  of  calcium  in  the  food  is  the  cause  of 
rickets  is  a  most  natural  assumption,  but  it  has  not  been  proved 
that  this  is  the  case.  Young  animals  fed  on  calcium-poor  foods  show, 
naturally  enough,  defective  development  of  the  bone,*'^  but  this  differs 
essentially  from  rickets  in  that  the  bone  formed  is  defective  chiefly 
in  amount  rather  than  in  quality  (Stoltzner).  Furthermore,  such 
"pseudo-rachitic  bone"  jDOSsesses'  a  marked  affinity  for  calcium  salts^ 
and  takes  them  up  as  soon  as  they  are  supplied  (Pfaundler).  In 
view  of  the  fact  that  rickets  is  not  solely  a  disease  of  bone  tissue,  but 
that  all  the  various  important  viscera,  as  well  as  the  muscles  and 
tendons,  show  pathological  changes,  it  seems  most  reasonable  that 
rickets  should  be  looked  upon  as  a  constitutional  disease,  in  which 
the  bone  changes  are  prominent  chiefly  because  the  disease  occurs  at  a 
time  when  the  bone  tissue  is  most  actively  forming  and  when  the 
other  organs  are  relatively  quite  completely  developed.  Stoltzner,*^ 
finding  evidence  that  rickets  does  not  depend  upon  either  lack  of 
calcium  in  the  food  or  deficient  absorption  of  calcium,  and  that  the 
blood  in  rickets  is  of  normal  alkalinity,  looks  upon  the  failure  of 
calcification  as  depending  upon  an  abnormality  in  the  calcified  bone 
tissue  itself.     He  finds  evidence  of  a  preliminary  alteration  in  normal 

44  Gassniann.  Zoit.  pliysiol.  Clicm.,  iniO   (70),  101. 

•»4<i  The  hones  and  muscles  in  Barlow's  disease  show  quite  the  same  deficiency 
in  calcium  as  in  rickets  (Balirdt  and  Kdelstein.  Zcit.  Kindcrheilk.,  1913   (9),  415). 

«La  Pediatria,  VIT,  497. 

^fia.Tahrb.  Kinderheilk..  1914  (79),  446.  Howland  and  Marriott  found  normal 
fipures   (Trans.  Amer.  Ped.  Soc,  vol.  28,  p.  202). 

■<<^  There  is  a  d<'crease  in  the  calcium  of  tlie  muscles  according  to  Aschenlieim 
and  Kaunihfimer    ( Monatschr.  f.   Fvinderheil..  1911    (10),  4.'i5). 

47Vcrliandl.  Dcut..  Path,  desell.,   190.")    (9).  248. 

47a  See  Weiser.   iJiochem.  Zeit.,  1914    (fifi),  95. 

48Jahrb.  f.  Kinderheilk.,  1899    (50),  208. 


coNCRErioxs  447 

osteoid  tissue  which  prepares  it  to  take  the  salts  out  of  the  bhxxl, 
and  Pfaundler  ^-  supports  this  view,  suggesting  that  this  prepara- 
tory change  in  the  osteoid  tissue  maj'  depend  upon  autolysis,  which 
is  perhaps  deticient  in  rickets.'"^ 

On  the  other  hand,  after  extensive  experimental  work,  Dibbelt  '^^ 
comes  to  the  conclusion  that  rickets  results  from  excessive  elimination 
of  calcium  into  the  intestine,  presumably  because  of  the  presence  of 
precipitating  substances  in  the  intestinal  contents,  such  as  P.Og  from 
casein.  Agreeing  with  Dibbelt  that  the  excessive  elimination  of  cal- 
cium is  chiefly  through  the  feces,  Schabad '"-  after  equally  extensive 
investigations,  believes  that  calcium  starvation  in  children,  from  de- 
fective absorption,  may  cause  at  least  a  pseudo-rickets,  indistinguish- 
able clinically  or  chemically  from  tme  rickets.  As  with  osteomalacia, 
attempts  have  been  made  to  associate  with  the  etiology  of  rickets  de- 
fects in  the  ductless  glands,  especially  the  adrenals,^^  thymus,*^^  and 
parathyroids,^*  but  as  yet  without  convincing  evidence."*'^ 

CONCRETIONS 

All  pathological  concretions  appear  to  be  laid  down  according  to  a 
definite  law.  There  must  first  be  a  nucleus  of  some  substance  differ- 
ent from  the  substance  that  is  to  be  deposited,  and  w^hicli  is  most 
frequently  a  mass  of  desquamated  cells,  but  may  consist  of  clumped 
bacteria,  masses  of  mucus,  precipitated  proteins,  or  a  foreign  body  of 
almost  any  sort.  Upon  this  nucleus  substances  crystallize  out  of 
solution,  much  as  cane-sugar  crystallizes  on  a  string  to  form  rock 
-•candy,  but  with  the  important  exception  that  among  the  crystals  is 
usually  deposited  more  or  less  mucin  or  other  organic  substance, 
M'hicli  forms  a  framework  in  w^hich  the  crystals  lie,  and  which  re- 
mains, if  the  crystals  are  dissolved  out,  as  a  more  or  less  perfect 
skeleton  of  the  concretion.  In  no  case  would  the  concretion  form 
were  it  not  that  the  solution  is  overcharged  with  some  substance,  but 
not  infrequently  it  is  the  presence  of  the  nucleus  that  leads  to  the 
precipitation  of  the  substance;  i.  e.,  the  nucleus  may  play  either  a 
primary  or  a  secondary  role.  AVith  few  exceptions,  the  dissolved 
substance  is  deposited  in  crystalline  form,  although  the  cn-stalline 
structure  may  in  time  partly  disappear  through  condensation  or 
through  filling  of  the  interstices  with  some  other  material.  Even  so 
structureless  a  substance  as  amyloid  may,  when  forming  concretions, 
appear  in  a  crystalline  form  (Ophiils).     The  structure  of  a  concre- 

50  See  also  Nathan.  :\red.  News,  1004   (84),  391. 

51  Articles  in  the  Arbeiten  a.  d.  Path.  Inst.  Tiibinofen,  Vols,  fi  and  7;  also  Verh. 
Dent.  Path.  Gesell.,  1010   (14),  204;  Miinch.  med.  Woch.,  1010  (57),  2121. 

52  Arch.  f.  Kinderhcilk..  1000  (52),  47;  1910  (53),  381;  1911  (54),  83; 
Fortschr.  Med.,   1010    (28),   1057. 

53  Stoeltzner,  Verh.  Dent.  Path.  Ges..  1009    (13),  20. 
5-4Erdhcini  et  al.  Frankfurter  Zeit.  Path.,  1911    (7),  178. 

54a  Concerning  the  chemical  changes  of  osteogenesis  imperfeeta  (congenital  fra- 
gility of  bones),  see  Schabad,  Zeit.  Kinderheilk.,  1914   (11),  230. 


448  CALCIFICATIOX,    COXVJi'KTWXS,    AXD    INCRUSTATIONS 

tiou  depends  upon  two  factors :  The  crystals  tend  to  be  deposited  at 
right  angles  to  the  surface,  and  thus  give  a  radiating  structure;  but 
the  rate  of  deposition  is  usuall}^  irregular,  and  during  the  periods  of 
quiescence  the  surface  tends  to  become  covered  witii  mucin  or  other 
organic  substances,  hence  we  also  get  a  concentric,  lami)iated  struc- 
ture. Frequently  both  of  these  lines  of  formation  are  easily  dis- 
cerned, but  either  one  or  the  other  may  become  obscured. 

Concretions  consist,  therefore,  of  mixtures  of  colloids  and  crystal- 
loids deposited  from  solutions  of  the  same  character,  and  hence  the 
application  of  the  principles  of  colloidal  chemistry  throws  much 
light  on  the  conditions  of  their  formation/''^  Colloidal  solutions 
hold  in  solution  greater  quantities  of  crystalloids  than  simple  solu- 
tions, for  the  reason  that  at  the  surface  of  each  colloidal  particle  there 
is  a  zone  in  which  the  crystalloids  are  more  concentrated  than  else- 
where, thus  permitting  more  crystalloids  to  be  dissolved  in  the  solvent 
between  the  colloidal  particles.  On  the  other  hand,  the  concentra- 
tion of  the  crystalloids  on  the  surface  of  the  colloidal  particles  causes 
the  colloids  to  serve  as  the  starting  point  of  precipitation  whenever 
the  crystalloids  are  in  excess.  When  the  crystalloid  goes  out  of 
solution,  therefore,  it  will  form  crystals  or  precipitates  which  are 
most  intimatel}^  associated  with  the  colloids,  as  we  see  when  uric  acid 
crystallizes  out  of  urine,  taking  with  it  the  colloidal  pigments  by 
which  it  is  adsorbed.  Or,  if  the  colloids  are  precipitated,  the  solvent 
power  of  the  solution  is  reduced,  and  the  crystalloids  will  deposit  in 
intimate  relation  to  the  colloids.  As  Schade  pointed  out,  if  a  colloid 
precipitates  in  an  irreversible  form  (e.  g.,  fibrin),  the  concretion  will 
be  permanent,  as  with  ordinary  concretions,  but  if  the  colloid  pre- 
cipitate is  reversible  the  mass  may  be  dissolved  again,  as  Avith  the 
precipitate  of  urates  in  the  tubules  of  the  infant's  kidney. 

BILIARY  CALCULI  55a 

As  may  be  judged  from  the  above  statements,  concretions  are  never 
composed  of  one  substance  in  a  pure  form,  but  usually  consist  of  a 
mixture  of  the  constituents  of  the  fluid  in  which  they  are  developed. 
This  is  particularly  true  of  gall-stones,  which  contain  in  greater  or 
less  (piantities  several  or  all  of  the  constituents  of  the  bile.  AVliile 
cholesterol  forms  the  greater  part  of  nearly  all  biliary  concretions,  and 
is  present  in  greater  or  less  amounts  in  all,  calcium  salts  of  the  bile- 
pigments  are  always  present;  usually  inorganic  salts  of  calcium  (car- 
bonate and  phosphate)  are  also  present,  as  well  as  small  amounts  of 
fats,  soaps,  lecithin,  mucus,  and  other  products,'''^''  and  occasionally 

55  See  Schade,  Miindi.  med.  Woch.,  1000  (.5(1).  :] -.  litll  (r)S),  r2■^■.  Zoit.  oxp. 
Path.,  1010  (8),  02:  also  Lichtwitz,  Krgcb.  inn.  MchI.,  I!)14  (i;{),  1;  also  his  nionn- 
grai)h  "Lecher  die  Hil(hin<r  dcr  ilani-  und  ( Jallenstcino,"  Spriiifier,  Berlin.   1!>14. 

5-.a  Bi),lio;,rrapliy  liy    Haciiicislcr,   Kr;,^^!).  inn.   Mod..  l!)l:?    (11),  1. 

r,ob  Fischer  and  Kosc  found  about  0.1  <im.  carotin  in  12S()  jiins.  >,'all  stones  from 
cattle.      (Zeit.  physiol.,  Cheni.,   li)13    (8S),  331.) 


JUIJAUY  CALCULI  449 

traces  of  copper/"'  iron,  and  iiiang-anese."  The  quantity  of  bile  salts, 
the  chief  constituent  of  the  bile,  is  usually  extremely  minute,  appar- 
ently only  so  much  as  may  percolate  into  the  crevices  of  the  concre- 
tion. However  many  stones  there  may  be  in  a  gall-bladder,  they 
usually  are  all  of  approximately  the  same  composition  and  structure. 

In  o-all-stones  from  the  domestic  animals  the  i)rop()rtion  of  inorganic 
salts  is  usually  much  higher  than  it  is  in  man. 

Naunyn  has  classified  gall-stones  according  to  their  composition,  as 
follows : 

1.  "Pure"  Cholesterol  Stones. — The  purity  is  only  relative,  since 
even  the  ])urost  always  contain  some  pigment  as  well  as  a  stroma 
and  a  nucleus;  but  the  amount  of  cholesterol  may  reach  98  per  cent., 
and  is  usually  over  90  per  cent.  Crystalline  structure  is  usually 
well  marked,  while  stratification  is  slight.  The  color  varies  from 
nearly  ]iure  white  to  yellow,  or  even  brown  on  the  surface. 

2.  Laminated  Cholesterol  Stones. — These  consist  of  about  75-90 
per  cent,  of  cholesterol,  and  differ  from  the  preceding  form  in  con- 
taining more  pigment,  which  is  deposited  in  layers  alternating  with 
the  white  layers  of  cholesterol.  The  pigrment  here,  as  in  all  other 
gall-stones,  consists  always  of  the  calcium  salts  of  the  pigments — 
not  of  pure  bilirubin  and  biliverdin  themselves.  Considerable  calcium 
carbonate  is  also  usually  present,  particularly  in  the  green  layers  of 
biliverdin-calcium. 

3.  Common  Gall-bladder  Stones. — The  composition  of  this  form  is 
but  little  different  from  the  above,  the  chief  difference  being  in  the 
structure.  They  present  externally  a  firmer  crust,  usually  distinctly 
laminated ;  in  the  center  is  a  softer  pigmented  nucleus  which  fre- 
quently shows  a  central  cavity  containing  fluid.  Such  calculi  are  not 
distinctly  crystalline  in  structure,  and  are  small,  seldom  larger  than 
a  cherry. 

4.  Mixed  Bilirubin-calcium  Calculi. — These  generally  occur  singly, 
but  sometimes  in  groups  of  three  or  four,  and  are  of  large  size. 
Although  the  chief  constituent  is  bilirubin-calcium,  there  is  always 
much  cholesterol,  often  over  25  per  cent.  Copper  and  traces  of  iron 
may  also  be  present.  Their  structure  is  laminated,  with  sometimes 
a  crystalline  cholesterol  nucleus. 

5.  "Pure"  Bilirubin-calcium  Calculi. — In  addition  to  the  chief  con- 
stituent, 'biliverdiii-calcmm,  hilifuscin  and  hilihumin  ^^  are  practically 

56  See  Mizokuclii,  Cent.  f.  Pathol.,  1912   (23),  3.37. 

57  Gall-stones  have  been  found  enclosing  droplets  of  merpury.  (Xaunyn. 
Frerichs. ) 

39  Biliverdin  differs  from  hiliruhin  in  containing-  one  more  atom  of  oxygen  in 
the  molecule,  and  it  is  easilv  formed  from  Ijilirubin — even  exposure  to  air  will 
slowly  bring  about  the  oxidation.  Bilifuscin  is  a  still  more  oxidized  deriva- 
tive— so  much  so  that  it  does  not  give  Gmelin's  reaction  (with  PTXO.  +  HNOj) 
for  bile-pigments.  Bilihumin  represents  the  most  oxidized  of  these  products, 
is  brown  in  color,  and  is  the  chief  constituent  of  the  residue  left  after  treating 
gall-stones  with  ether,  alcohol,  and  chloroform  to  dissolve  out  the  cholesterol. 

29 


450  CALCIFICATIOW    ('<)\Cin:TI()\S,    AM)    IWhTSTATlOXS 

always  present.  liilihumiii  is  at  times  tlie  chief  iiiKi'eilient.  and  niay 
form  over  half  of  the  substance;  bilicijanin  is  rarely  present.  There 
is  always  some  cholesterol,  but  sometimes  onlj^  traces.  These  calculi 
are  small,  from  the  size  of  a  grain  of  sand  to  that  of  a  pea,  and  they 
occur  in  two  distinct  forms.  One  form  is  of  wax-like  consistence;  the 
other  is  harder.  steel-<>Tay  or  black  in  color,  with  a  metallic  luster. 
Pure  bi]irul)iii  and  hiliverdin,  not  coinhiiicd  with  calcium,  are  prac- 
tically never  ]i resent  in  concretions. 

6.  Rarer  Forms. —  (a)  AniorpJwus  (okI  incomplete]])  cri/stdlJine  cho- 
lesterol gravel.  Cholesterol  externally  giving'  them  a  pearly  luster: 
pigment  in  the  center. 

(6)  Calcareous  Stones. — Consist  chiefly  of  a  mixture  of  calcium 
carbonate  and  bilirubin-calcium.  Calcium  carbonate  may  occur 
either  as  a  superficial  crust,  or  as  small  masses  within  an  ordinary 
calculus;  calcium  sulphate  and  phosphate  occur  rarely  in  traces. 
Stones  consisting  mainly  of  calcium  carbonate  are  extremely  rare  in 
man,  but  more  frecjuent  in  cattle  and  other  herbivora,  in  which  all 
forms  of  concretions  contain  much  calcium,  either  combined  with 
pigment  or  as  carbonate  and  phosphate.  A  calcium  oxalate  gall-stone 
has  also  been  described.*^" 

(c)  Concretions  ivith  included  bodies,  and  conglomerate  stones. 

(d)  Casts  of  Bile-ducts. — Occur  particularly  in  cattle,  and  consist 
chiefly  of  bilirubin-calcium.     Rarely  and  imperfectly  formed  in  man. 

Formation  of  GalUstones. — We  owe  much  of  our  present  under- 
standing of  the  chemistry  and  pathology  of  the  formation  of  gall- 
stones to  Naunyn  "^  and  his  pupils.  Former  observers,  having 
learned  that  bile  normally  contains  cholesterol  (Hammarsten  found 
from  0.06-0.16  per  cent,  in  human  bile),  sought  the  cause  of  gall- 
stones in  either  an  increased  elimination  of  cholesterol  by  the  liver,  or 
a  decrease  in  the  power  of  the  bile  to  hold  the  cholesterol  in  solution. 
Thus  Frerichs,  finding  that  the  presence  of  large  amounts  of  bile 
salts  and  an  alkaline  reaction  favored  the  solution  of  cholestei-ol,  im- 
agined that  a  diminution  of  either  bile  salts  or  alkalinity  led  to  the 
precipitation  of  the  cholesterol.  Naunyn  and  his  pupil's,  however, 
observing  that  the  amount  of  cholesterol  present  in  the  bile  does 
not  depend  upon  the  amount  taken  in  the  food  or  tlie  amount  present 
in  the  blood,  and  that  it  did  not  vary  in  disease,  exce]it  when  gall- 
stones were  present,  concluded  lliat  the  cholesterol  of  the  bile  is 
neither  a  product  of  geiu'ral  metabolism  nor  a  specific  secretion-prod- 
uct of  the  liver.  Finding  that  ])us  and  the  secretions  from  inflamed 
iiiiii'ous  iiK'inbranes  (bronchitis)  contained  as  much  cholesterol  as  did 
iioniial  bile,  and  often  more,  they  concluded  that  the  chief  source  of 

'^'1  Mniit  lain-.    Hull.  sci.   |pliaiiiiaf(il..   \'ol.    IS,  p.   111. 

'•'  All  l-^iifrlisli  translation  of  tliis  classic  work,  by  .\.  K.  (iarrod,  lias  Iuhmi  |)u1)- 
lislicd  by  the  Sydenham  Society,  ISiXi,  vol.  l.'iS.  "Nfoio  recently  excejition  has 
been  biken  to  certain  of  Nanii.\ii's  \  icws,  cs|)ccia]ly  by  AsdiolV  and  nacnicister. 
"Cholelithiasis,"  Custav  Fisclicr,  .Iciia.   l!l()!l. 


lill.lMH  rM.ci  /.I  451 

eliolestorol  in  ^all-stoiu'  roniiatioii  was  fi-oiii  tin-  (lc;:riK'ratiii^  and 
desquamated  epithelial  cells  of  the  jiall-hladdcr  and  l)ilr  tracts.  This 
idea  was  supported  by  the  larjie  amount  of  cholesterol  found  in  the 
coutents  of  tiall-l)ladders  shut  off  from  the  conunon  duct,  and  by  the 
formation  of  (jall-stones  in  such  isolated  <iall-bladders.  P^urther  evi- 
dence has  since  been  brought  forward  in  favor  of  this  same  view,^- 
until  it  has  been  generally  accepted  as  correct,  although  there  are  some 
who,  tinding  no  abundance  of  cholesterol  in  the  wall  of  the  gall 
bladder,  do  not  accej)t  this  origin."-' 

On  the  basis  of  this  hypothesis  the  ordinarv  steps  in  the  formation 
of  a  cholesterol  concretion  are  as  follows :  Some  injury  to  the  mucous 
membrane  of  the  bile  tracts  is  the  starting-point ;  this  injury  is 
usually  pi'oduced  by  infection,  the  colon  and  typhoid  bacilli  being 
the  most  common  organisms  in  this  process. ''^  Through  the  degener- 
ation of  the  epithelial  cells  an  excess  of  cholesterol  is  formed,  while  at 
the  same  time  the  desquamated  cells  and  clumped  bacteria  offer 
suitable  nuclei  upon  Avhich  the  cholesterol  begins  to  crystallize  out. 
Apparently  after  the  calculi  have  reached  a  certain  size  they  cause 
sufficient  mechanical  injury  to  keep  up  the  cell  degeneration  and 
cholesterol  formation,  even  after  the  infection  has  subsided.  A  cer- 
tain amount  of  infection  and  inflammation  is  a  favoring  condition, 
however,  for  Harley  and  Barratt ''"'  found  that  fragments  of  chol- 
esterol calculi  introduced  aseptically  into  the  gall-bladders  of  dogs  were 
slowly  dissolved  and  disappeared,  but  this  was  prevented  by  infecting 
the  gall-bladder  with  B.  coli.  According  to  Naunyn's  investigations, 
it  is  not  an  alteration  in  the  composition  of  the  bile,  as  formed  in 
the  liver,  which  causes  the  precipitation  of  cholesterol,  but  rather  the 
presence  of  the  nidus,  and  the  production  of  large  quantities  of 
cholesterol  in  immediate  proximity  to  this  nidus,  that  determines  the 
formation  of  a  concretion.  In  case  the  bile  stagnates  in  the  gall- 
bladder, the  cholesterol  that  is  being  constantly  formed  by  the  normal 
disintegration  of  surface  epithelium  accumulates,  until,  even  without 
infection,   there   forms   a    sediment   of   soft   yellowish   and   brownish 

02  Thus  ^Yakeman  I  quoted  by  Herter,  Trans.  Congress  Amer.  Pliysicians,  100.3 
(6),  158:  excellent  resume)  was  able  to  cause  an  increase  in  tlie  cliolesterol  of 
the  bile  in  the  gail-ijiadder  of  dogs  by  injecting  into  it  HgCL,  phenol,  or  ricin. 
At  first  the  cliolesterol  seems  to  be  contained  largely  in  tlie  degenerating  des(|ua- 
mated  cells.  Also  the  interesting  case  of  a  cholesterol  calculus  in  a  pvosalpinx, 
described  by  Thies  (Arb.  Path.  Inst.  Tiibingen,  1908  ((i),  422),  shows  the  pos- 
sibility of  an  inflammatory  origin  for  such  concretions,  and  independent  of  bile. 

'■>3  Aschotr,  Miinch.  med.' Woch..  1906  (53).  1847  and  1913  (00),  1753;  Laroche 
and  Flandin,  Compt.  Rend.  Soc.  P.iol..  1912   (721.  (i60. 

04  See  Cushing  (Jt)hns  Hopkins  Hosp.  Bull.,  1899  (10).  l()(i),  who  produced 
gall-stones  experimentally  by  injecting  typhoid  bacilli  into  the  circulation  after 
injuring  the  gall-bladder.  Literature  (m  the  relation  of  bacteria  to  gall-stones 
given  by  Fnnke,  Proc.  Path.  Soc,  Philadelphia.  1908  (11),  17:  also  see  Uosenow 
who  finds  that  streptococci  are  often  res!)onsible  (Jour.  Infect.  Dis.,  191fl  (19), 
527). 

«3Jour.  of  Physiol.,  1903  (29),  341;  see  also  Hansemann.  Vircli.  Arch.,  1913 
(212),  139. 


452  CALCIFICATION,    CONCRETIOXS,    .l.VZ)    IXCRl'STATIOyS 

masses,  consisting  chiefly  of  cholesterol  and  bilirubin-calcium.  From 
this  material  calculi  may  eventually  form,  and  by  their  irritation  lead 
to  further  formation  of  cholesterol  and  increased  o:rowth.''*'^  But 
bacterioloo-ical  studies  indicate  that  generally  an  infectious  influence 
is  present  in  cholelithiasis,  and  bacilli  may  be  found  alive  in  gall-stones 
for  remarkably  long  periods. 

Recent  applications  of  colloidal  chemistry'  add  much  to  our  under- 
standing of  gall-stone  formation.  Thus,  Lichtwitz  points  out  that 
the  colloids  of  normal  bile,  all  of  which  are  electro-negative,  may  be 
precipitated  by  positive  serum  colloids  coming  from  the  blood  when 
the  gall-bladder  is  inflamed ;  hence  we  get  a  precipitate  of  cholesterol, 
bilirubin  and  proteins.  Wlien  the  colloids  are  thus  thrown  down  the 
solvent  power  of  the  bile  for  the  alkali  earths  it  contains  is  decreased, 
and  so  calcium  or  magnesium  are  added  to  the  mixture.  Cholesterol 
is  in  solution  in  the  bile  as  an  emulsion  colloid,  and  when  stagnation 
of  the  bile  leads  to  absorption  or  disintegration  of  the  cholates  and 
fats  which  keep  it  in  solution,  the  droplets  become  confluent,  and 
then  ciystallization  takes  place  (Schade)  with  formation  of  sphero- 
liths,  and  eventually  a  crystalline  cholesterol  calculus.  If  cA'en  the 
slightest  pressure  is  brought  to  bear  on  the  myelin-like  masses  before 
they  crystallize,  however,  they  will  be  pressed  into  scales,  and  the 
common  laminated  structure  results ;  hence  crystalline  calculi  are 
single,  while  multiple  gall-stones  are  laminated,  with  perhaps  partial 
crystallization  between  the  lamellae.  Also  when  the  gall-stones  result 
from  inflammation,  and  there  is  much  serum  colloid  present,  the 
stones  are  lamellated  because  these  colloids  deposit  in  that  form  (e.  g., 
corpora  amylaeea  and  other  protein  concretions).  These  considera- 
tions explain  the  formation  of  gall-stones  in  the  gall-bladder  from 
either  inflammation,  or  stagnation  without  inflammation. 

AschofiP  and  Bacmeister,*'"  however,  hold  that  the  usual  series  of 
events  in  the  fonnation  of  gall-stones  is  first  the  formation  of  a  pure 
cholesterol  stone  without  inflammatory  cause,  because  of  actual  in- 
creased excretion  of  cholesterol  by  the  liver,  because  of  cholester- 
olemia,  or  because  of  resorption  of  solvent  substances  from  stagnating 
bile;  these  primary  cholesterol  stones  then  cause  inflammation  and 
occlusion,  leading  to  the  formation  of  the  common  mixed  stones. 
Bacmeister  ascribes  more  importance  to  calcium  than  do  most  other 
investigators,  while  Kuru  ^^  states  that  fibrin  is  usually  present. 

More  recent  studies  of  the  cholesterol  content  of  the  blood  and  bile 
also  have  reacted  against  the  concept  that  all  the  cliolesterol  of  gall- 
stones comes  from  the  wall  of  the  bile  tract  througli  inflammatory 
changes.     It  has  been  found  that  patients  with  gall-stones  often  show 

"^a  Concerninfj  the  stnicturo  of  frail  stonos  sec  Kibhort.  VirchowV   Aroli..   IHlo 
(220),  20. 
nn  Zioplor's  Beitr.,  lOOR   (44),  .'i2S. 
«7Virc]iow's  Arch.,  1912    (210),  4.1.'^. 


BILIAKY  CALCULI  453 

a  hypereliolesterolemia  (Ilenes""'')  ;  that  pregnancy,  which  seems  to  be 
a  predisposing  cause  of  cholelithiasis,  is  accompanied  by  hyperchol- 
esterolemia; that  with  hypercholesterolemia  there  is  an  increased  out- 
put of  cholesterol  in  the  bile,  and  that  experimental  hypercholesterole- 
mia may  lead  to  the  formation  of  gall-stones  without  evident  infection 
of  the  bile  tracts  (Dewey  °'^).     As  far  as  the  existing  evidence  permits 
one  to  draw  conclusions,  it  would  seeui  probable  that  both  local  and 
systemic  conditions  are  of  importance  in  gall-stone  formation.     Ap- 
parently, gall-stones  may  form  from  cholesterol  derived  from  the  in- 
flamed bile  tract  walls,  independent  of  the  amount  of  cholesterol  pres- 
ent in  the  bile ;  but  presumably  they  may  derive  part  if  not  all  the 
cholesterol  from  the  bile  in  some  cases.     In  either  event,  a  hyper- 
cholesterolemia will  favor  their  formation,  and  hence  any  given  con- 
dition of  injury  to  the  gall  bladder  will  more  often  give  rise  to  con- 
cretions in  persons  with   a  high  cholesterol   content   in   the   blood. 
Changes  in  the  bile  itself  may  be  produced  by  disease  of  the  liver  that 
will  alter  its  composition  in  such  a  way  that  its  capacity  to  sustain 
cholesterol  in  solution  or  suspension  will  be  lowered,"*^  and  this  fac- 
tor also  cannot  be  dismissed  as  without  importance ;  transient  thicken- 
ing of  the  bile,  such  as  may  occur  in  any  febrile  disease,  may  also  very 
possibly  initiate  precipitation  and  stone  formation.*^'*^ 

It  w-as  formerly  supposed  that  the  calcium-pigment  concretions 
were  produced  by  the  presence  of  excessive  calcium  in  the  bile,  de- 
rived particularly  from  lime-laden  drinking-water,  but  it  has  been 
demonstrated  that  increase  of  calcium  in  the  food  does  not  cause  an 
increase  in  the  amount  in  the  bile.  Furthermore,  on  concentrating 
bile,  which  contains  both  bilirubin  and  calcium,  the  free  bilirabiu 
separates  out  and  not  the  calcium  compound  of  bilirubin;  and  also, 
Naunyn  found  that  the  bile  salts  prevent  precipitation  of  calcium- 
bilirubin,  even  when  calcium  salts  are  added  in  considerable  amounts. 
Apparently  it  is  the  presence  of  positively  charged  protein  substances 
that  leads  to  the  precipitation  of  this  electro-negative  substance  from 
bile,  and  hence  the  formation  of  pigment  calculi  is  also  favored  or 
initiated  by  inflammation  of  the  bile  tracts,  particularly  as  most  of 
the  calcium  salts  seem  to  come  from  the  mucous  membrane ;  ^^  later, 
as  we  have  seen,  these  pigment  concretions  often  become  covered  with 
cholesterol  derived  from  the  injured  epithelium,  and  the  common  mixed 
calculi  are  then  formed.  In  view  of  the  fact  that  much  of  the  pig- 
ment in  these  calculi  is  composed  of  the  oxidation  products  of  bili- 

67a  Surg.,  Gvn.  and  Obst..  lOlf,   (2.3),  91. 

sTbArch.  Int.  Med.,  1016  (17),  757;  see  also  Aoyama,  Deut.  Zeit.  Chir.,  1914 
(1.32),  234. 

cTcSee  D'Amato,  Biochem.  Zeit.,  1915    (69),  .353. 

67dSee  Rovsing.  Hospitalstidende,  1915   (58),  249. 

(■■8  This  commonly-held  view  is  denied  by  Lichtwitz  and  Bock  (Deut.  med. 
Woch.,  1915  (41),  1215),  who  found  the  calcium  content  of  bile  from  fistulas  to 
be  from  65-84  mg.  per  liter,  and  in  l)hidder  bile  to  vary  from  85  to  325  mg.,  but 
not  according  to  the  presence  or  absence  of  inflammation. 


454        CALCJFicAriox,  coxcnirr/oxs.    \\i>  ixchTsTAriows 

rubin,  especially  billhunilii,  it  is  p()ssil)l('  that  oxidatiou  processes  in 
the  stagnating  bile  are  im])ortant  causes  of  the  precipitation;  Naunyn 
suggests  that  bacteria  may  be  the  cause  of  the  oxidation.  Pigment 
calculi  are  particularly  important  as  the  starting-point  of  the  larger 
mixed  calculi.  It  is  possible,  Naunyn  believes,  for  the  pigment  to  be 
later  gradually  replaced  hy  cholesterol. 

URINARY  CALCULI  >■•■> 

These  differ  from  the  l)ile  concretions  in  two  imi)ortant  respects: 
First,  there  is  no  evidence  that  any  considerable  part  of  tlieir  con- 
stituents may  come  from  the  walls  of  the  cavities  that  contain  them; 
they  are  usually  deposited  on  account  of  an  over-saturation  of  the 
urine,  or  on  account  of  a  change  in  composition  of  the  urine,  which 
renders  them  insoluble.  Second,  the  composition  of  urinary  calculi  is 
usually  less  mixed  than  that  of  biliary  calculi,  although  seldom,  if  ever, 
is  it  pure.  Thus,  Finsterer  found  but  six  concretions  composed  of 
only  one  substance,  in  a  collection  of  114  calculi.  As  with  the  bile,  the 
chief  constituent  of  the  urine  (urea)  is  so  soluble  that  it  never  forms 
concretions,  but  only  the  less  soluble  minor  constituents  are  thrown 
down.  For  the  formation  of  calculi,  however,  it  is  not  sufficient  to 
have  merely  an  excess  of  a  substance  in  the  urine,  for  we  may  have 
deposition  of  urates,  phosphates,  or  uric  acid  in  simple  crystalline 
form  without  the  formation  of  calculi.  A  nucleus  of  some  sort  is 
present  as  well  as  a  binding  substance,''^  which  is  often  mucus  derived 
from  the  walls  of  the  passages,  although  the  center  of  the  concretion 
most  often  consists  of  uric  acid  or  urates. 

Although  the  amount  of  colloidal  material  in  urine  is  relatively 
small,  yet  it  undoubtedly  plays  an  important  part  in  maintaining  in 
solution  the  less  soluble  crystalloids,  which  are  especially  the  urates 
and  calcium  oxalate.  Normal  urine  contains  no  colloids  which  form 
irreversible  gels,  and  hence  ordinary  deposits  can  be  readily  dissolved, 
but  in  inflammatory  conditions  there  appears  fibrinogen  which  readily 
forms  the  irreversible  fibrin,  and  conditions  thus  become  favorable 
for  the  formation  of  concretions  of  any  crystalloid  with  which  the 
urine  m'dy  be  saturated  or  over-saturated  at  the  time  (Schadeh 
Possibly  other  colloids  may  play  a  similar  role.  Aschoff'  and  Kleiu- 
schmidt "'  hold  that  most  urinary  calculi  begin  as  primai-y  calculi, 
formed  independent  of  inflammation  from  excess  of  the  nmin  con- 
stituent (uric  acid,  oxalates,  xanthine,  but  chiefly  ammonium  urate)  : 
this  calculus  foniis  the  ciystalline  nucleus  of  tlie  laminated  second- 
ary deposits  of  other  sul)stances,  chiefly  uric  acid,  oxahites  and  phos- 

•■'!•  fipneral  liihiiofrrapliy  ;iivoii  liy  Fiiistoror.  Dcut.  Zcit.  kliii.  Ch'w..  I'.iOi;  (SO). 
414;  and  Liclitwitz.'^^'' 

70  Hippocrates  a])preciatpil  tlie  existence  and  iniijoiiaiicc  <it"  the  mucoid  liiii(lin<r 
substance  in  urinarv  eoneretions  (Sdiepelniann.  I?erl.  kiin.  Wocli.,  IHII  (4S), 
525 ) . 

VI  "Die  iiarnsteine,"  lU-ilin,  .luiius  Spriufior,   IDll. 


/  /.'/A   I  AM     (A  LCI  I.I  455 

])Jiatos.  all  hciii^-  deposited  williout  iiiHamuiatioii.  The  iidlaiuiiiatory 
formations  consist  cliictlN  of  ammonio-magnesiuiii  |)lins|)liate  and 
aninioiiium  nriite,  usually  deposited  on  a  foreign  body  or  a  primary 
ealeulus.  The  extensive  study  of  tiie  microscopic  structure  of  urin- 
ary calculi  by  Shattock,'-  shows  also  that  a  nucleus  of  cells  or  other 
organic  material  is,  at  least  in  uric  acid  calculi,  extremely  rare,  the 
center  being  almost  always  a  primary  crystalline  deposit  from  a 
supersaturated  solution. 

Calculi  formed  because  of  changes  in  the  uriiuiry  composition  in- 
dependent of  evident  infection  are  often  called  "primary."  in  con- 
tradistiiu'tiou  to  those  arising  from  changes  in  composition  brought 
about  b\  infection  and  ammoniacal  decomposition.  Because  of  the 
injury  ])r()duced  by  a  primary  calculus,  infection  frequently  results, 
and  then  tlie  prinuiry  calculus  may  become  the  nucleus  of  a  secondar^^ 
calculus ;  indeed,  on  account  of  the  change  of  reaction,  the  crystalloids 
of  the  primary  calculus  may  be  dissolved  out,  and  their  place  taken  by 
the  secondary  deposit  (metamorphosed  calculi).  In  structure  urin- 
ary calculi  usually  show  both  radiating  and  concentric  lines  of  forma- 
tion, and  when  the  chief  constituents  are  dissolved  away,  an  organic 
framewoi"k  remains.  They  are  generally  classified  according  to  their 
])rominent  component,  as  follows: 

Uric=Acid  Calculi. — Uric  acid  is  but  slightly  soluble,  only  one  part 
dissolving  in  39,480  of  pure  water  at  18°,  and  it  is  even  less  soluble 
in  the  presence  of  acids.'-"^  The  presence  of  sodium  diphosphate  in  the 
solution  makes  it  much  more  soluble,  and  various  organic  bodies  also 
favor  its  solution,  among  them  being  the  urinary  pigments.  As  can 
be  seen,  the  maintenance  of  uric  acid  in  solution  is  by  a  small  margin, 
even  in  normal  conditions;  hence  the  mere  cooling  of  the  urine  fre- 
quently suffices  to  cause  an  abundant  deposition  of  uric  acid  combined 
with  pigment,  as  the  familiar  "brick-dust"  deposit.  The  formation 
of  uric-acid  calculi  is,  therefore,  not  only  a  question  of  the  amount  of 
uric  acid  in  the  urine,  but  depends  even  more  upon  the  amount  of  the 
substances  that  hold  it  in  solution,  and  as  both  these  factors  are  sub- 
ject to  wide  variations  under  both  physiological  and  pathological  con- 
ditions, uric  acid  and  urates  are  common  in  urinary  conci'etions. 

The  older  literature  indicates  that  the  most  common  calculus  is  of 
this  nature,  but  a  number  of  recent  analyses  indicate  that  the  im- 
portance of  uric  acid  and  urates  has  been  overestimated.  On  the  con- 
trary, this  material  rarely  forms  a  considerable  part 'of  the  calculi, 
but  is  usually  present  in  greater  or  less  amount  in  most  or  all  urinary 
calculi  (Kahn ).'•"'     It  is  probable,  however,  that  uric  acid  is  imi)ortaiit 

"2  Proc.  Roy.  Soc.  Med.,  Path.  Sec,   1911    (4),  110. 

'2a  Concerning  soliihilitv  of  uric  acid  in  urine  see  Ifaskins.  .lour.  P.iol.  (liem., 
191(5   (-26).  205. 

'-Arch.  Int.  Med.,  1913  (11).  92;  review  of  literature.  Rosenhloom,  (.Tour. 
Amer.  ^led.  Assoc,  1915  (05),  101)  found  but  two  uric  acid  stones  of  twenty-six 
analyzed. 


456  CALCIFICATION,    COXCKETIO\>S,    AXD    IXCRU^TATIONS 

as  furnishing  the  primary  nucleus  of  calculi  of  preponderatingly  cal- 
careous or  mixed  composition.  Apparently  there  are  marked  differ- 
ences in  the  prevailing  composition  of  calculi  in  different  countries ;  in 
China,  for  example,  Pfister  "'^  found  eleven  of  twelve  calculi  composed 
of  uric  acid. 

Uric  acid  is  eliminated  combined  chiefly  with  sodium,  potassium^ 
and  ammonium ;  according  to  some  authors,  as  a  biurate,  according  to 
others,  as  a  quadriurate.  If  the  urine  is  excessively  acid,  it  con- 
tains much  acid  phosphates,  which  withdraw  part  of  the  bases  from 
the  uric  acid,  and  this,  when  free,  crystallizes  out  if  in  excess.  Hence 
tlie  foruuition  of  uric-acid  concretions  is  favored  by  high  acidity  of 
the  urine,  by  concentration  of  the  urine,  or  by  an  increased  elimina- 
tion of  the  uric  acid.  The  last  may  result  from  excessive  nuclein- 
rich  food,  or  from  excessive  catabolism  of  the  tissue  nucleoproteins 
(e.  g.,  leucocytosis  from  inflammatory  diseases  or  leukemia),  which 
conditions  are  also  usually  associated  with  an  increased  urinary  acid- 
ity. (The  chemistry  of  uric  acid  is  discussed  more  fully  in  the  chap- 
ter on  ''Gout,"  Chap,  xxi.) 

Uric-acid  calculi  are  formed  chiefly  in  the  pelvis  of  the  kidney,  but 
many  pass  into  the  bladder.  They  are  quite  hard,  and  yellow  or 
reddish-yellow  in  color,  because  of  the  presence  of  vrochrome  and 
K'rohilin,  the  former  of  which  seems  to  be  chemically  combined  and 
the  latter  but  physically,  since  it  can  be  washed  out  with  v/ater. 
Uraerythrin  or  uromelanin  (a  decomposition  product  of  urochrome) 
may  also  be  present.  Not  infrequently  calcium  oxalate  is  present, 
sometimes  in  considerable  quantities.  Other  urinary  constituents  may 
be  present  in  small  amounts.  In  case  the  calculus  enters  the  urinary 
bladder  it  may  set  up  irritation  leading  to  infection;  the  urine  then 
becoming  alkaline,  calcium  and  ammonio-magnesium  phosphate  will 
be  deposited  upon  the  surface,  and  the  uric  acid  will  be  more  or 
less  dissolved  out  and  replaced  by  the  phosphates  (metamorpho- 
sis). 

Urate  calculi  occur  chiefly  in  new-born  or  young  infants,  and  rarely 
in  adults.  In  the  young  thoy  are  related  to,  and  may  originate  in, 
the  deposits  of  urates  in  the  pyramids  of  the  kidney  (tlie  so-called 
urate  or  uric-acid  "infarcts"),  which  have  been  supposed  to  result 
from  the  decomposition  of  the  nucleoproteins  of  the  nucleated  fetal 
red  corpuscles.  (See  "Uric  Acid,"  Chap,  xxi.)  The  concretions  are 
composed  chiefly  of  either  ammonium  or  sodium  urate,  but  potassium 
and  even  calcium  and  magnesium  urate  may  be  admixed.  Their 
genesis  in  the  young  probably  depends  upon  injury  to  epithelium  by 
the  excessive  urates  of  tlie  "infarcts,"  whicli  affords  a  suitable 
nucleus  for  their  start;  their  growth  depends  chiefly  upon  the  con- 
centration of  the  infant's  urine.  In  adults  they  may  arise  secondary 
to  an  ammoniacal  decomposition  of  the  urine.     Urate  concretions  are 

73aZeit.  Urol.,  1M13   (7),  945. 


LV.'/A   l/.T  VALVULI  457 

not  coiiimuu ;  they  are  generally  rather  soft,  and  often  mueh  colored 
by  pigments. 

Calcium  oxalate  calculi  are,  according  to  recent  observers,'^  the 
most  oonunon  urinai-y  concretions.'^  Often  they  show  admixtures  of 
urates  or  uric  acid,  which  latter  frc(iuently  constitutes  the  nucleus,  and 
when  urinary  infection  occurs  they  may  in  turn  serve  as  the  nucleus 
to  phosphatic  deposits.  On  account  of  the  hardness  and  roughness  of 
these  stones  they  frequently  cause  bleeding,  which  may  result  in  their 
being  very  dark  in  color  and  containing  blood-pigment.  They  are 
usually  first  formed  in  the  pelvis  of  the  kidney,  and  arise  chiefly  in 
persons  excreting  excessive  quantities  of  oxalic  acid.  Normally  but 
about  0.02-0.05  gram  of  oxalic  acid  is  eliminated  daily  in  the  urine, 
apparently  all  as  calcium  oxalate,  which  is  kept  in  solution  by  the  acid 
phosphates.  The  amount  may  be  increased  by  certain  foods  rich  in 
oxalates,  particularly  rhubarb,  grapes,  spinach,  etc. ;  also  probably 
by  gastric  fermentation."^  Oxalic  acid  may  possibly  be  formed  from 
uric  acid,  and  perhaps  also  from  the  carbohydrate  group  of  pro- 
teins,'*^ and  it  is  possible  that  abnormally  large  amounts  arise  from 
these  sources  under  pathological  conditions.  During  bacterial  de- 
composition of  the  urine  oxalic  acid  miay  be  formed  from  uric  acid 
(Austin).^' 

Phosphate  calculi  are  formed  as  a  result  of  decomposition  of  the 
urine,  with  formation  of  ammonia  from  the  urea.  In  the  ammoniacal 
solution  thus  formed  the  magnesium  is  precipitated  as  NH^MgPO^, 
the  calcium  as  Ca3(P04)„,  and  calcium  oxalate  and  ammonium  urate 
are  also  thrown  down,  so  that  the  concretions  consist  of  a  mixture  of 
these  substances,  the  magnesium  salt  being  the  most  abundant.  In 
none  does  one  substance  occur  in  a  pure  state.  Pigments  of  various 
kinds,  and  more  or  less  mucus  or  other  organic  constituents  of  the 
framework  are  also  present.  Phosphate  calculi  are  the  typical  "sec- 
ondary" concretions,  and  they  are  formed  usually  in  the  bladder  as  a 
consequence  of  cystitis,  but  may  be  formed  in  the  renal  pelvis  or  in 
the  urethra.  In  some  cases  the  salts  are  precipitated  in  such  large 
quantities  that  they  form  great  masses  of  a  sediment  which  does  not 
aggregate  into  concretions.  Occasionally  stones  consisting  princi- 
pally of  Ca.,(POJ.  or  CaHPO^  are  formed,  but  these  are  rarities.  As 
the  calcium  taken  in  the  food  is  chiefly  eliminated  in  the  feces,  the 
amount  in  the  urine  does  not  vary  directly  with  the  amount  in  the 
food,  and  the  formation  of  phosphatic  concretions  is  always  a  matter 
or  urinaiy  reaction  and  not  of  diet.'*     As  these  stones  fuse  to  a  black, 

7*  Concerning  their  structure  see  Fowler,  Johns  Hopkins  Hospital  Reports,  1906 
(13)..';07. 

"5  Baldwin,  Jour.  Exp.  Med.,  1900   (5),  27. 

-«  See  Austin,  Boston  Med.  and  Surg.  Journal,  1901  (145),  181.  Contradicted 
by  Wegrzynowski.  Zeit.  phvsiol.  Cliem..  1913    (83),  112. 

TT.Tour.  Med.  Research.  1906   (15),  314. 

78  Under  the  name  "struvit  stone,"  Poninier  (Verh.  deut.  Path.  Gesell..  1905 
(9),  28)  describes  a  urinary  calculus  composed  of  very  pure  ammonio-magnesium 


458         cALcii'icATiow   c()\('i>'i:ti(>\s,  am)   /xrh'i  statioxs 

enaiiu'l-like  mass  under  the  l)l()W-i)ipe,  they  have  been  called  "fusible 
calculi." 

Calcium  carbonate  calculi  are  formed  freciuently  in  herbivora, 
but  tliey  are  xcry  rare  in  the  \irinary  passages  of  man,  althoufi-h  oe- 
currinw  elsewhere  in  the  body  not  infrequently.  Occasionally  these 
are  soft  and  chalky,  but  if  well  crystallized,  they  are  the  hardest  of 
concretions. 

Cystine  calculi  '"  are  rare  but  very  interesting  formations.     Cys- 

S-CII(NIL)-C'0011 

tine     I  is    important    as    the    sulphur-containing    por- 

S-CH(XIL)-COOH 

tion  of  the  protein  molecule.  Under  normal  conditions  all  the  cystine 
taken  in  food  is  completely  oxidized  and  none  (or  uncertain  traces) 
appears  in  the  urine.  In  certain  individuals  the  urine  contains  con- 
siderable quantities  of  cystine  constantly  {(jjstiuuria,  see  Chap,  xix), 
and  occasionallj"  in  these  cases  soft  concretions  of  nearly  pure  cystine 
are  formed  in  the  urinary  passages.  Cystine  calculi  may  reach  the 
size  of  a  hen's  egg,  are  crystalline  in  structure,  and  in  the  urine  of 
such  patients  the  characteristic  hexagonal  crystals  may  usually  be 
found.     The  cystine  of  calculi  is  identical  with  that  from  proteins.^" 

Xanthine  Calculi. — Xanthine  is  the  most  abundant  of  the  purine 
bases  normally  present  in  urine,  but  the  total  amount  is  extremely 
small.  Like  uric  acid,  it  fluctuates  in  amount  according  to  the 
amount  of  destruction  of  nucleoproteins,  either  of  the  food  or  of  the 
tissues.  Concretions  consisting  chiefly  of  xanthine,  which  is  often 
mixed  with  uric  acid,  are  extremely  rare,  but  a  few  isolated  si)eci- 
mens  having  been  described.  Rosenbloom  could  collect  but  six  cases 
in  the  literature,  adding  one  himself. ^^ 

Indigo  calculi,  derived  from  the  indican  of  the  urine  through  oxi- 
dation, have  also  been  described  a  few  times. 

Urostealith  calculi,  composed  of  fatty  matter,  have  been  occasion- 
ally ol)served.  Although  some  of  the  concretions  described  under 
this  head  have  really  represented  foreign  bodies  introduced  through 
the  urethra  (e.  g.,  Kruckenberg 's  concretion  of  paraffin  from  a  bou- 
gie), yet  true  fat  concretions  do  occur.  The  origin  of  the  fat  in  tliese 
stealiths  is  unknown;  possibly  it  comes  from  degenerated  epithelium. 
TTorbaczewski  ^'  analyzed  such  a  specimen  which  had  the  following 
percentage  composition : 

Water 2..'> 

Tnoriranio  matt«M"     ....  OS 

Orfjaiiic    matter    (cliiclly    ])riitciii  ) 11.7 

Fatty    acids .')]..") 

Nculral    fal :?:5..-, 

Cliolcstcnil  traces 

])li()S|)lia1e,  foriiiiii^  tlic  liard,  rlioinliic  crystals  known  to  iiiiiicralouisis  as 
"struvit."  'I'liis  is  an  exaini)lc  of  a  l)lios|)Iiate  stone  formed  independent  of  am- 
inf)niaea]  deeom]H>8iti()ii,  a  raic  occurrence. 

'»  T>iteratnre  concerniiij,'  cystine,  see  Kricdmann,  ImltcIi.  der  IMi\siol.,  ]'J()2  (iK 
1.'):    Marriott   and  Wolf.  Am."  .lour.   Med.  Sci.,   1!1()(>    (i:?!).   \'M. 

'^'1  .M.dcriialden.   Zcit.   iilivsioi.   Clicm..    1!»07    (51),  :{91. 

«i  X.   V.   Med.  .loiir.,  .lati.   Ill,   lUlf). 


/  A'/N    I  A')     CM. CI  I.I  459 

The  fatty  acids  coiisisti'd  of  stearic,  j)aliiiitic,  and  pi'ohalily  inyi-istic 
aeid. 

Cholesterol  calculi  have  been  found  in  the  urinary  bladder  in  a 
few  instances,  the  cause  being  unknown.  Horbaczewski ''-  descril)es 
one  weij2;hiiig  25.4  ••■rains,  found  in  a  patient  who  had  i)i-eviously  had 
cystine  calculi;  it  contained  1)5.87  per  cent,  of  cholesterol  and  but  0.55 
per  cent,  of  inorganic  inatei'ial.  (iall-stones  have  been  known  to 
enter  the  urinary  bladder  through  a  fistula  between  the  gall-bladdei- 
and  urinary  bladder."" 

Fibrin  "calculi,"  formed  from  blood-clots,  often  moi"e  or  less  im- 
pregnated with  urinary  salts,  have  occasionally  been  observed.  Other 
proteins  may  also  form  similar  calculi.*^ 

General  Properties  of  Urinary  Concretions/"* — The  hardness  de- 
])ends  partly  upon  the  chemical  composition  of  the  calculus,  hut  more 
upon  the  rate  and  condition  of  formation  (Rowlands,  Kahn).  I'nder 
comparable  conditions  it  is  said  that  those  composed  of  amorphous 
phosphates  are  the  softest ;  next  come  those  with  some  admixture  of 
crystalline  phosphates.  IVate  concretions  are  harder  than  these,  but 
are  still  softer  than  the  uric  acid  and  crystalline  phosphate  calculi. 
Oxalates  are  usually  the  hardest,  except  for  the  rare  crystallized 
calcium  carbonate  stones.  Cystine  and  amorphous  concretions  can 
be  scratched  with  the  finger-nail,  while  even  the  hardest  varieties  of 
calculi  can  be  scratched  with  a  wire  nail.  Genersich  -"  gives  the 
following  degrees  of  hardness  for  different  calculi:  Cholesterol,  1.5- 
1.6;  ammonium  urate,  2.5;  soft  phosphate  (^Ig),  2.6;  hard  phos- 
phate (Ca).  2.75:  uric-acid  stones  (also  salivary  and  prostatic  calculi, 
atheromatous  patches,  and  phleboliths),  2.9;  calcium  oxalate  (also 
rhinoliths  and  lung  stones),  3.3-3.5;  calcium  carbonate  stones  of 
herbivora,  4.5.  But  the  hardness  or  gross  appearance  of  a  urinary 
calculus  give  little  or  no  indication  of  its  chemical  composition. 

The  rate  of  growth  also  varies  according  to  composition,  but  is,  of 
course,  mucli  modified  by  other  factors.  Oxalate  and  urate  stones 
grow  most  slowly,  phosphate  stones  most  rapidly.  A  urate  stone  has 
been  known  to  increase  by  about  two  ounces  during  seven  and  one 
half  years,  while  a  catheter  fragment  or  other  foreign  body  may  be- 
come covered  with  a  crust  several  millimeters  thick  in  a  few  weeks."" 

Spontaneous  disintegration  of  urinary  concretions  is  limited  almost 
solely  to  calculi  composed  entirely  or  largely  of  uric  acid.  Out  of  121 
cases  collected  by  Englisch,*^  in  all  but  7  this  was  the  case,  these  being 

82  Zeit.  physiol.  Cliem.,  1804    (IS),  Xir^. 

83  See  Finsterer,  Deut.  Zeit.  kliii.  Chir.,  lOm;    ISO),  4-i(). 

S4  See  ]\Iorawitz  and  Adrian,  Mitt.  Orcnz.  Med.  u.  Chir.,  l!li)7    i  17).  .■)7!). 

f<">  Systems  for  i)rocediire  in  detenninin'/  tlie  nature  of  luinarv  ealeuli  are  given 
by  Hammarsten    (Text-l)ooi<  of  Phvsiol.  C'liem.)    and  bv  Smith    (  Reference  Hand- 
book of  Med.  Sci.). 
■     sfi  Virehow's  Arch..  180.3   (131),  185. 

s"  Zuckerkandl.  Xothnasjel's  System,  vol.  10,  pt.  2,  p.  220. 

ssAroh.  klin.  Chir.,  100.>    (76),  Or,l    (elaborate  review). 


460  CALCIFICATION,    CONCRETIONS,    AND    INCRUSTATIONS 

composed  of  calcium  and  niagnesiuin  phosphate  (5),  or  calcium  phos- 
phate or  carbonate  (1  each).  The  tlisintegration  is  brouglit  about 
through  solution  of  the  binding  substance  and  mechanical  shattering 
of  the  stone  into  fragments.  This  occurs  but  rarely,  Bastos  ®*^  esti- 
mating that  perhaps  one  calculus  in  ten  thousand  undergoes  disinte- 
gration. 

CORPORA  AMYLACEA  ^^ 

In  the  case  of  these  widely-spread  concentric  bodies  we  find  the 
name  misleading,  for  the  bodies  are  not  a  form  of  animal  starch,  as 
was  suggested  by  their  laminated  structure  and  iodin  reaction,  nor 
are  they  so  closely  related  to  amyloid  material  as  the  name  implies. 
Different  authors  disagree  decidedly  concerning  the  staining  reactions 
of  these  bodies,  but  it  may  be  said  that  the  reactions  are  extremely 
inconstant.  Sometimes  the  corpora  are  stained  bluish  or  green  with 
iodin,  sometimes  brown,  often  little  at  all ;  occasionally  they  react 
partly  with  methyl-violet,  but  more  often  they  do  not ;  sometimes  por- 
tions of  one  body  react  one  way,  while  the  remainder  behaves  differ- 
ently. Seldom  if  ever  do  the  ordinary  concretions  of  the  prostate 
give  all  the  amyloid  reactions  characteristically,  but  the  corpora 
amylacea  of  the  lungs  are  much  more  likely  to  do  so  (Stumpf).''°  It 
seems  improbable  that  these  bodies,  which  occur  in  the  prostate  of 
every  adult,  can  be  the  same  as  the  amyloid,  which  is  seldom  observed 
except  as  the  result  of  serious  processes  of  tissue  destruction.  Accord- 
ing to  their  structure  they  obey  the  usual  laws  of  the  formation  of 
concretions,  having  a  central  nucleus  and  a  structural  framework  of 
different  composition  from  the  chief  substance.  It  seems  most  prob- 
able that  they  should  be  interpreted  as  simple  concretions  of  protein 
nature,  Avhich  form  under  certain  conditions  when  a  nucleus  of  some 
sort  (usually  pigment,  degenerated  cells,  or  inorganic  crystals)  exists 
in  a  stagnating,  protein-rich  fluid.  At  times  the  resulting  concretion 
may  be  of  such  a  physical  nature  that  it  absorbs  iodin  readily  (just 
as  they  often  show  a  marked  absorption-affinity  for  pigments),  and 
occasionally  it  may  react  metachromatically  with  methyl-violet,  pos- 
sibly because  of  the  presence  of  chondroitin-sulphuric  acid  derived 
from  the  mucin  of  the  cavities  where  the  concretions  form,  but  per- 
haps for  some  other  unknown  reasons.  Occasionally  pure  amyloid 
may  form  in  the  tissues  typically  concentric  (or  even  crystalline) 
bodies,  as  in  Ophlil's  case,  but  this  is  the  exception.  It  seems  prob- 
able that   corpora  amylacea  are   usually   protein   concretions,"^   and 

88a  Folia  Tirol.,  101.'?    (8),  SI. 

80  Gonoral  literature,  Posncr,  Zcit.  kliii.  Mod.,  ISSO  (Ifi),  144:  Luharscli, 
Erpcl).  allfr.  Patliol..  1894  (L).  ISO;  Opliiils,  .Tour.  Exp.  :\lo(l..  1900  (5).  Ill; 
Nunf)I<a\va.  Vircliow'a  Arch.,  1909    (19(5),  221;   Briitt,  ibid.,   1912    (207),  412. 

00  Vircliow's  Arch.,  1910  (202),  1.34. 

01  Ramsdon'.'^  olisorvations  (Pror.  Royal  Soc,  190.3  (72),  156).  on  tlic  procipi- 
tation  of  j)roleiiis  l)y  tlio  action  of  surface  contact  may  liave  some  bcarinir  on 
the  formation  of  such  protein  concretions. 


CONCRETIONS  461 

neither   amyloid   nor   animal   stareli.     Those    i'ormed   in    the   central 
nervous  system  may  be  of  myelin  or  nenrojjlia  origin. "- 

The  small  amount  of  material  available  prevents  an  accurate 
analysis  of  the  eor})ora  amylacea;  it  is  known  that  they  are  very  in- 
soluble in  water,  acids,  alkalies,  etc.,  behaving  like  coagulated  protein 
in  this  respect.  p]ven  hot  concentrated  nitric  acid  will  not  dissolve 
them,  according  to  Posner.  This  author  considers  lecithin  and  cho- 
lesterol to  be  important  constituents,  and  by  Ciaccio's  staining 
method  lipoids  can  be  found  in  prostatic  corpora  amylacea."^  How- 
ever, it  is  said  by  Bjorling  ^*  that  the  ordinary  hyaline  and  granular 
corpora  do  not  contain  fats  or  lipoids,  but  that  a  certain  class  of 
"lipoid"  prostatic  concretions  contain  many  granules  of  this  nature. 
The  corpora  amylacea  of  the  lateral  ventricles  seem  to  consist  chiefly 
of  calcium  salts  deposited  in  a  concentric  arrangement  through  the 
medium  of  an  organic  basis.  Posner  considers  that  the  presence  of 
lecithin  in  prostatic  corpora  prevents  their  calcification,  although 
this  change  occasionally  does  occur. 

OTHER  LESS  COMMON  CONCRETIONS 

Pancreatic  Calculi."' — The  cause  of  the  formation  of  stones  in  the 
pancreatic  duct  is  not  definitely  known,  but  apparently  infection  is 
the  most  important  factor,  since  simple  experimental  stasis  will  not 
cause  their  formation."^  The  calculi  consist  usually  of  a  mixture  of 
calcium  phosphate  and  carbonate,  associated  Avith  more  or  less  organic 
matter,  including  frequently  cholesterol,  but  all  the  usual  products  of 
proteolysis  may  be  present  because  of  the  presence  of  trypsin.  Oc- 
casionally the  calculi  consist  chiefly  of  calcium  carbonate,  which  may 
be  almost  pure."'"'  Shattock  °'^  has  observed  a  pancreatic  concretion 
composed  of  calcium  oxalate.  Sodium  phosphate  and  chloride,  mag- 
nesium phosphate,  and  proteins  have  also  been  found  in  these  concre- 
tions. Taylor  "®  describes  a  pancreatic  concretion  containing,  accord- 
ing to  the  analyst,  chiefly  silicate  (!),  a  finding  difficult  to  under- 
stand or  accept. 

Baldoni  "^  found,  on  analysis  of  a  stone  weighing  3.1  grams,  the 
following  percentage  composition : 

Water .1.44 

Ash 12.f)7 

Proteins  .3.40 

Free    fatty    acids 1,3. ,39 

Neutral  fatty  acids 12.40 

92  See  Lafora,  Virchow's  Arch.,  1911    f20r>),  29,5. 
03  Posner,  Zeit.  f.  ITrologie,   1911    (,5),  722. 
^i  Tbid.,  1912   (6),  30. 

95  Literature  by  Scheunert  and  Berpliolz,  Zeit.  plivsiol.  Chom..   1907    (-52),  33S. 

96  See  Lazarus,' Zeit.  klin.  Med..  1901   (,51),  530.     Literatin-e. 
9fia  Rosenthal,  Arch.  f.  Verdauunuskr.,  1914    (20).  G19. 

97  Brit.  Med.  .Tour.,  1896    (i),  1034. 

98  Lancet,  Dec.  18,  1909. 

99  Schmidt's  Jahrb.,  1900   (268),  210. 


462         cALciFKwnow   coscinrnoxs.  .\\i>  i\Ch'rsT.[Ti<)\^< 

Cholesterol 7.69 

Pifrnients    and    soaj) 40.01 

Uiuletormiiied 6.01 

rsually.  liowc'ver,  pancreas  stones  consist  chiefly  of  inorganic  sub- 
stances. .Johnson  and  Wollaston  report  analyses  of  two  stones,  one 
containing  72.30  per  cent,  calcium  phosphate  and  but  8.80  per  cent. 
organic  matter;  the  other  91.65  per  cent,  calcium  carbonate,  4.15  per 
cent,  magnesium  carbonate,  and  but  3  per  cent,  organic  matter. 
Legrand  ^  found  only  0.7  per  cent,  organic  matter  i)i  another  concre- 
tion which  contained  93.1  per  cent,  calcium  carbonate.  Pancreatic 
juice,  being  strongly  alkaline,  can  hold  but  a  small  quantity  of  calcium 
salts  in  solution  (normally  but  0.22  part  per  thousand — C.  Schmidt)  ; 
presumably  the  little  normally  present  is  held  in  the  form  of  a  colloidal 
suspension  by  the  proteins.  Possibly  when  stasis  occurs,  digestion  of 
the  proteins  leads  to  the  precipitation  of  the  calcium  salts,  or,  more 
probably,  the  excessive  calcium  is  largely  derived  from  the  exudate 
from  the  inflamed  ducts,  as  seems  to  be  the  case  with  the  calcium  of 
biliary  calculi. 

Salivary  Calculi.- — These  have  a  similar  composition,  in  the  main, 
to  the  concretions  of  the  pancreatic  duct,  except  that  they  generally 
contain  more  organic  matter,  resembling  in  this  respect  the  "tar- 
tar" of  the  teeth.  Bessanez  found  in  one  81.3  per  cent,  of  calcium 
carbonate  and  4.1  per  cent,  of  calcium  phosphate,  whereas  in  an- 
other the  carbonate  was  but  2  per  cent,  and  the  phosphate  75  per 
cent.  Potties  has  described  a  calculus  with  a  central  portion  com- 
posed chiefly  of  uric  acid  and  a  peripheral  portion  containing  69  per 
cent,  of  calcium  pliospliate  and  20.1  per  cent,  of  calcium  carbonate. 
Harlay  ^  found  in  one  specimen  15.9  per  cent,  organic  matter,  75.3 
per  cent,  calcium  phosphate,  6.1  per  cent,  calcium  carbonate.  Ro- 
berg  believes  that  bacteria  alone  do  not  usually  cause  salivary  calculi 
to  form,  but  that  a  foreign  body  entering  the  duct  is  the  chief  factor. 
Increased  alkalinity  nuiy  also  favor  ]irecipitation  of  calcium  from  the 
saliva.  In  Roberg's  case  of  sialolitliiasis  the  saliva  was  of  normal 
composition. 

Intestinal  Concretions. — These  always  have  a  nuclens  of  some 
indigestible  foreign  su])stance.  most  often  hair,  but  sometimes  eellu- 
lose  structures  or  solid  indigestible  j^articles,  including  gall-stones. 
fruit-stones,  bone,  etc.  The  bulk  of  the  eoiu'retions  is  usually  made 
U]i  chiefly  of  ammonio-magnesium  ))hosj)hate,  with  some  calcium 
])hos])hate,  carbonate,  and  sulphate,  |)rotein  matter,  and  occasionally 
calcium  and  magnesium  soa])s.  Two  iulestiual  coiu'retions  analyzed 
by  Scliuberg  ^  had  the  following  percentage  com{)osition  when  dried: 

1  .Idiir.    I'lianii.  ct  Cliiiii..    I'.IOI    (14),  21. 

-  I.iteratiire,  see   KoIht'.',   Annals  of   Siiruerw    l!>(»4    (."!!»),  (Itl!). 

a.Ioiir.    I'liarm.  et   Cliiin.,    l'.Mt:{    (IS),    11. 

■•  \'irclio\v's   .Arcii.,    ISSii    (!l(l|,   ~:i. 


IXTEiiTl\AL  COS VRET loss  M).\ 

Ainnionio-iiiayiicsiuin   pliospliate      ....      ■)7.1  (i;5.!t 

Calcium  phospliatf '"i-T  23.S 

Calcium  carbonate 4.(5 

Calcium  sulpiiate -J.*'  ".7 

Alcoiiol-ctlicr  extract l.'.t  n.s 

Other  orgajiic  substances -lo  ti.n 

111  L'uuntries  wliere  oatmeal  is  largely  eaten,  intestinal  eoneretions 
are  not  infre<inent ;  they  contain  calcium  and  magnesium  phosphate, 
about  70  per  cent.;  oatmeal  bran,  15-18  per  cent.;  soaps  and  fats, 
about  10  per  cent.  (Hammarsten).  Occasionally  concretions  con- 
sisting: largely  of  fats  and  soaps  are  found,  and  after  taking  large 
doses  of  olive  oil  masses  of  solidified  oil  may  be  passed  that  are  read- 
ily mistaken  for  softened  gall-stones,  for  the  removal  of  whicli  the 
oil  is  usually  given.  The  "fecal  stones"  found  in  appendices  often 
show  the  structure  of  calculi,  and,  unlike  other  enteroliths,  consist 
less  of  ammonio-magnesium  phosphate  than  of  calcium  salts;''  soaps 
may  be  important  constituents.® 

Bezoar  stones  are  intestinal  concretions  probably  coming  from 
Capra  aegagrus  and  Atitelope  dorcas.  One  variety  consists  chiefly 
of  lithofellic  acid,  C.,^,'H.^^,0^,  which  is  related  to  cholalic  acid,  and 
gives  an  aromatic  odor  when  heated.  The  other  variety  ("false 
bezoars")  does  not  give  the  aromatic  odor,  and  consists  chiefly  of 
ellagic  acid,  C^Ji^fi^^,  a  derivative  of  gallic  acid,  and,  therefore,  prob- 
ably derived  from  the  tannin  of  the  food  of  the  antelopes. 

Intestinal  "sand"  occurs  as  (1)  "false  sand,"  consisting  of  parti- 
cles of  indigestible  food,  such  as  the  sclerenchymatous  particles  in 
the  flesh  of  pears  and  bananas;  '  and  (2)  trne  sand,  consisting  largely 
of  inorganic  material,  and  formed,  according  to  Duckworth  and  Gar- 
rod,^  in  the  upper  part  of  the  large  intestine.  Analyses  of  speci- 
mens b}'  Garrod  showed  the  following  composition : 

Water        ....      12.4  f    calcium    oxide .54.98 

Orofanic   material            2C).29                        )  jiliosphorus     pentoxidc     .      .      .  42. .35 

Inorganic  material    .      G1.31  containing!     carbon    dioxide 2.20 

I  traces   of   ^Ig,    Fe,    etc.    .                     0.47 

Analyses  by  other  observers  have  given  similar  results,  the  absence  of 
the  large  proportion  of  magnesimn  found  in  larger  concretions  being 
striking. 

The  color  is  usually  brown,  due  chiefly  to  urobilin,  unaltered  bile- 
pigments  being  scanty. 

Preputial  concretions  sometimes  form  beneath  a  prepuce  that  can- 
not he  retracted,  through  deposition  of  urinary  salts  on  and  in  the 
accumulated  smegma.^  The  composition  is,  therefore,  verj^  mixed, 
and  consists  of  an  organic  base  containing  much  cholesterol,  fats,  and 

5  Harlav,  Jour,  pharm.  et  chim.,  1010   (2),  43.3. 

6  Williams.  Biocliem.  Jour.,  1007    (2),  395. 

TMyer  and  Cook.  Amer.  Jour.  Med.  Sci.,  1000    (137),  383. 
s  Lancet,   1902    (i).  053.     Full  resume  and  literature. 
9  See  Zeller,  Arch.  klin.  Chir.,  1890   (41),  240. 


464 


CALCIFICATION,    CONCRETIOyS,    AND    IXCRUSTATIOyS 


soaps,  incrusted  with  inorganic  substances,  of  which  ammonio-magne- 

sium  phosphate  and  calcium  phospliate  are  usually  the  most  abundant. 

Prostatic  concretions  originate  in  the  corpora  amylacea  through 

growth  accretion  of  inorganic  salts,  until  they  may  reach  considerable 

size.     Stern  "  gives  the  following  results  of  analysis  of  such  a  prostatic 

stone : 

Water         8.0 

Organic   matter  15-8 

Lime 37.64 

Magnesia 2.38 

Soda 1-76 

Potash 0.5 

Phosphoric  acid 33.77 

Iron trace 

Lung  stones.' 1 — These  may  be  formed  in  the  bronchi,  through  ac- 
cretion about  an  inorganic  nucleus,  similar  to  the  formation  of  cal- 
culi in  other  epithelial-lined  passages ;  or  they  may  consist  of  calcified 
areas  of  lung  tissue  or  peribronchial  glands,  which  have  been  se- 
questrated through  suppuration  and  have  entered  the  bronchi.  In 
the  latter  case,  the  calculi  present  the  usual  composition  of  patho- 
logical calcified  areas.  That  the  expectorated  stones  frequently  rep- 
resent calcified  tubercles  is  shown  by  Stern  ^^  and  by  Biirgi,^^  who 
demonstrated  tubercle  bacilli  in  decalcified  lung  stones.  The  follow- 
ing percentage  figures  are  taken  from  Ott :  ^- 

Calcium    phosphate 52.0  72.8 

]\Iagnesiimi    phosphate 
Magnesium    carbonate 


1.0 


2.0 

Calcium    carbonate 13.0 

Fat   and    cholesterol 24.0 

Other    organic    substances 4.0 


6.0 

7.0 

10.0 


Rhinoliths "  are  formed  about  nasal  secretions,  blood-clots,  and 
most  frequently  about  foreign  bodies.  They  therefore  contain  much 
organic  substance  in  addition  to  the  inorganic  salts  deposited  upon 
them.  Berlioz  ^^  gives  the  following  table  from  the  analysis  of  four 
specimens : 


Weight  of  specimens,  grams... 

1 
3.75 

1.34 

3 

0.63 

4 
0.95 

Water 

Organic   matter    .... 
Calcium   phosjjhate    . 
^Magnesium    phosphate    . 
Calcium    carbonate    . 
Traces  of  iron      .... 

5.80 
Ki.tiU 
02. 02 

5.08 

10.50 

Doubtful. 

5.10 

18.2(1 

(j().(il 

(i.28 

0.81 

Distinct. 

4.00 
l(i.()i) 

(;i.4(» 

14.(i7 
Doubtful. 

6.90 

18.10 

47. ()3 

G.OS 

20.(i!» 

Distinct. 

loAmer.  Jour.  Med.  Sci.,  1903   (126),  281. 

11  Literature.  Poulalion.  Thesis,  Paris,  1891;  Stern,  Deut.  med.  Woch.,  1904 
(30),  1414.  Biirgi  (Deut.  med.  Woch.,  1906  (32),  798);  Gerhartz  and  Strigel, 
Beitr.  z.  klin.  Tuberc,  1908    (10),  33. 

i2"Chem.  Path,  der  Tuberc."   1903,  p.  92. 

13  Literature,  Schc))pegrell,  Jour.  Amer.  Med.  Assoc,  1S9C  (20),  874;  Gerber, 
Deut.  med.  Woch.,   1892    (18),   1165. 

"Jour.  Pharm.  et  Chim.,  1891    (23),  447. 


]'\/:t  MOxohOMOSis  465 

Tonsillar  concretions  consist  chiefly  of  carbonate  and  phosphate 
of  calcium  deposited  upon  tlie  inspissated  secretions  and  desciuaniated 
cells  of  the  tonsillar  crypts.^"'  According  to  some  authors,  leptothrix 
threads  frequently  form  the  nucleus  of  the  concretions. 

Cutaneous  concretions  are  occasionally  observed,  located  chiefly 
in  tlie  suhcutaiieous  tissue,  often  occurring  multiple.  The  origin  is 
possibly  in  dihited  sebaceous  glands  with  retained  secretions.  Unna 
considers  that  calcium  soaps  are  formed  as  a  first  step,  but  an  analy- 
sis of  such  material  by  Harley  ^^  showed  87.2  per  cent,  of  ash,  12.8 
per  cent,  organic  matter,  0.9  per  cent,  of  fat;  calcium  phosphate  con- 
stituted 65.2  per  cent.,  and  calcium  carbonate  16.4  per  cent.  Gas- 
card  ^'  found  in  similar  material  23.4  per  cent,  organic  matter,  and 
of  the  inorganic  matter,  91.1  per  cent,  was  calcium  phosphate,  and 
8.9  per  cent,  calcium  carbonate. 

Gouty  deposits  observed  in  the  subcutaneous  tissues,  as  well  as 
along  the  tendons,  articular  cartilages,  etc.,  consist  usually  of  nearly 
pure  biurate  of  sodium  and  potassium.  Ebstein  and  Sprague  ^*  found 
the  composition  of  such  material  to  be  as  follows : 

Uric    acid 59.70 

Tissue  organic   matter 27.88 

Sodium  oxide n.30 

Potassium    oxide 2.95 

Calcium   oxide 0.17 

MgO,  Fe,  P2O,,  S traces 

After  a  time,  however,  calcium  salts  may  be  deposited,  and  Dunin  ^^ 
has  observed  deposits  resembling  gouty  tophi  that  were  merely  cal- 
cium salts. 

PNEUMONOKONIOSIS 

In  a  number  of  cases  of  the  different  forms  of  this  condition  quan- 
titative analyses  have  been  made,  which  may  be  briefly  discussed  as 
follows :  Not  only  does  the  lung  of  every  adult  contain  considerable 
amounts  of  coal-pigment  stored  up  in  the  connective  tissues  (and  also 
in  the  peribronchial  glands),  but  also,  which  is  perhaps  less  generally 
appreciated,  considerable  quantities  of  silicates  are  also  present  (chal- 
icosis)  from  inhaled  dust.  AVoskressensky  -'^  found  silicates  in  all  of 
54  lungs  examined,  except  two  from  infants.  The  lungs  of  individ- 
uals whose  occupations  do  not  expose  them  especially  to  dust  inhala- 
tion contain  increasing  amounts  of  silicates  in  direct  proportion  to 
age ;  the  silicates  constitute  then  from  3.5  to  10  per  cent,  of  the  total 
ash  of  the  lungs.     There  is  always  a  larger  proportion  of  silicates 

i5:McCarthy,  Brit.  Med.  Jour..  Oct.  28.  1911. 
16  Jour.  Pharm.  et  Chim.,  1903   (18),  9. 
IT  Ihid..  1900   (12).  262. 
isVirchow's  Arch.,   1891    {\2r^),  207. 

i9:Mitt.  Grenzpeb.  Med.  u.  Chir.,   190.",    (14).  4r>l :   also  Kahn.  .\rch.  Int.  Med., 
1913    (11).  92.  and  M.  B.  Schmidt,  Deut.  med.  Woch.,  1913    (39),  59. 
20  Cent.  f.  Path..  1S98    (9),  296. 
30 


466  CALCIFICATIOX,    COXCRETIOXS,    AND    INCRUSTATIOXS 

in  the  peribrone-liial  glands  than  in  the  lungs,  constituting  from  6  to 
36  per  cent,  of  the  ash,  corresponding  with  Arnold's  observation  that 
in  gold-beaters  the  glands  contain  more  metal  than  the  lungs.  In 
stone-workers  Schmidt  found  a  higher  proportion  of  SiO.  in  the  lungs 
than  in  the  glands.  In  normal  adults  the  amount  of  coal-pigment 
is  greater  than  the  amount  of  silicates;  in  children  the  reverse  is  the 
case. 

Thorel  -^  reports  that  the  lungs  of  a  worker  in  soapstone  contained 
3.25  per  cent,  of  ash,  including  2.43  per  cent,  of  soapstone. 

In  siderosis  iron  has  been  found  in  the  lungs  in  proportions  varying 
from  0.5  per  cent,  to  7.9  per  cent,  of  the  dry  weight,  the  last  amount 
having  been  found  by  Langguth  "  in  the  lungs  of  an  iron  miner,  which 
contained  also  11.92  per  cent,  of  SiOo. 

An  analysis  of  a  lung  from  a  knife-grinder  is  reported  by  Iloden- 
pyl,-^  Avhich  gave  the  following  results :  Total  weight  of  dried  and 
powdered  lung,  48.1009  grams ;  total  solids,  44.7986 ;  ether-soluble  sub- 
stance, 14.6017.  Composition  of  the  ether-soluble  substance :  free 
fatty  acids,  7.498;  neutral  fats,  4.044;  cholesterol,  3.037.  Proteins, 
15.4759;  charcoal  (total  carbon  less  protein  carbon),  7.198;  ash, 
4.2903.  The  composition  of  the  ash.  (in  grams)  was  as  follows:  KoO, 
0.2167;  Xa..O,  0.3523;  CaO,  0.0965;  Fe,0;,„  0.0879;  AUO„  1.4628; 
SO,,  0.0704;  P,0-„  0.9565;  SiO„  1.2043.  The  amount  of  emery,  rep- 
resented by  the  oxides  of  aluminum  and  silicon  made  up  more  than 
one-half  of  the  ash,  and  the  iron  constituted  about  one-fourth.  The 
man  had  worked  at  the  trade  of  knife-grinder  for  about  fifteen  years. 

]\IcCrae  -^  has  analyzed  the  lungs  of  six  gold  mine  workers,  in  South 
Africa,  finding  from  9  to  21.7  grams  of  ash  per  lung,  of  which  29  to 
48  per  cent,  was  silica;  aluminum  was  also  high,  and  an  increased 
P2O..,  content  was  ascribed  to  the  accompanying  fibrosis.  Klotz  -■' 
found  from  1.2  to  5.3  grams  of  free  carbon  in  each  lung,  of  dwellers 
of  Pittsburg,  as  contrasted  with  0.145  and  0.405  grams  found  in  the 
lungs  of  residents  of  Ann  Arbor.  Hirsch  -•'  analyzed  four  average 
Chicago  lungs,  finding  in  grams  per  lung: 

I 

Carbon 2.72 

Silica O.IS 

Calcium    O.xidc        .  .      0.45 

21  Ziegler's  Beitr.,  1896   (20),  85. 
22Dcut.  Arch.  klin.  Med.,  1895    (55),  255. 

23  Medical  Record.  1890    (56),  942. 

24  "The  Ash  of  Silicolic  T.niifrs."  John  McCrae,  Johannesburg,   1914. 
2''' Anicr.  .T(nir.  Piibl.  Tb'altli,  1914    (4),  887.     General  review  on  antliracosis 
20  Jour.  Amer    Med    Assoc,  191()    (66),  9.50. 


TT 

Til 

IV 

0.71 

1.20 

0.19 

0.28 

0.69 

0.04 

0.12 

0.02 

0.05 

CHAPTER    XVI 
PATHOLOGICAL  PIGMENTATION  ' 

MELANIN  - 

Melanin  occurs  normally  as  the  coloring-matter  of  hair,  of  the 
choroid  of  the  eye,  of  the  skin,  in  the  pigment  matter  of  many  lower 
animals,  and  most  strikingly  as  a  defensive  substance  in  the  "ink'' 
ejected  by  squids  to  render  themselves  invisible  in  the  water.  Path- 
ologically melanin  occurs  chiefly  as  the  result  of  an  excessive  pro- 
duction of  this  pigment  by  cells  normally  forming  it,  as  in  freckles, 
melanotic  tumors,  and  Addison's  disease  (probably).  Cells  that  do 
not  normally  form  melanin  probably  do  not  acquire  this  power  in 
pathological  conditions.-"  Pathological  failure  to  form  melanin  is  also 
observed,  as  in  skin  formed  in  the  healing  of  wounds  and  after 
syphilitic  lesions;  or  in  albinism,  in  which  the  failure  to  form  me- 
lanin may  be  attributed  to  hereditary  influences.^  The  function  of 
melanin  is  evidently  that  of  protection  from  light  rays,  and  Young  ^'' 
has  found  that  isolated  melanin  from  human  skin  absorbs  violet  and 
ultra-violet  rays.  Probably  this  protection  is  responsible,  at  least  in 
part,  for  the  relative  infrequency  of  skin  cancers  in  the  colored 
races.^'' 

Melanin  seems  always  to  be  produced  through  metabolic  activity 
of  specialized  cells.  The  idea,  which  was  formerly  advanced,  that 
it  is  derived  from  hemoglobin  as  a  product  of  disintegration,  seems 
to  have  failed  entirely  of  substantiation.  In  malaria  we  frequently 
find  a  diffuse  pigmentation  of  the  skin  of  such  a  nature  as  to  suggest 
strongly  a  melanin  formation,  and  this  has  been  cited  as  an  example 
of  the  production  of  melanin  from  hemoglobin.  Carbone  has  proved, 
however,  that  this  malarial  pigment  is  derived  from  hematin.  The 
amount  of  iron  contained  in  melanin  has  been  much  investigated,  as 

1  Literature  bv  Oberndorfer,  Ergebnisse  Pathol.,  1908  (12),  460,  and  Hueck, 
Zieofler's  Beitr.,  1012    (54),  (58. 

2  Literature  and  resume  given  by  v.  Fiirth.  Cent.  f.  Pathol.,  1004  (15),  617; 
Handb.  d.  Biochem..   1,  742. 

2a  The  pigment  of  the  so-called  "melanosis"  of  the  large  intestine  is  neither 
true  melanin  nor  ordinary  "waste"  pigment  (Henschen  and  Bergstrand,  Ziegler's 
Beitr.,  1013    (56),  10.3). 

3  Gortner  holds  that  dominant  whites  are  due  to  the  presence  of  antioxidase. 
while  regressive  whites  have  neither  the  power  to  form  pigments  nor  to  inhibit 
their  formation    (Amer.  Naturalist,  1010    (44),  497). 

3a  Biochem.  Jour.,  1014   (8),  460. 

3b  However.  Hanawa  found  white  areas  in  sl;in  less  aflfocted  by  cliemical  irri- 
tants and  infections  than  dark  areas.      (Dermatol.  Zeit.,  1013    (20),  761.) 

467 


468  PATHOLOGICAL  PIGMENTATION 

bearing  upon  the  question  as  to  whether  the  melanin  is  derived  from 
hemoglobin  or  not,  and  the  results  obtained  by  the  best  methods  indi- 
cate that  the  amount  of  iron  present  is  usually  extremely  small,  and 
often  it  is  entirely  absent;  furthermore,  the  presence  of  iron  is  no 
proof  that  the  pigment  is  derived  from  hemoglobin,  since  other  iron- 
protein  compounds  undoubtedly  exist, — especially  nucleoproteins,  and 
chemical  examination  shows  that  melanin  does  not  contain  hemopyr- 
role  groups.* 

Composition  of  Melanin. — The  elementary  composition  of  differ- 
ent specimens  of  melanin  examined  by  various  observers  has  been 
found  to  vary  greatly.  This  probably  depends  on  three  factors: 
First,  it  is  extremely  difficult  to  obtain  melanin  in  a  pure  condition; 
second,  the  process  of  purification  requires  the  action  of  strong  acids 
and  alkalies,  which  undoubtedly  modify  the  composition  of  the  mel- 
anin; thirdly,  melanin  is  probably  not  a  single  substance  of  definite 
composition,  but  includes  several  related  but  different  bodies.  The 
values  found  vary  for  carbon  from  48.95  to  60.02  per  cent. ;  for  hy- 
drogen from  3.05  to  7.57  per  cent. ;  for  nitrogen,  8.1  to  13.77  per 
cent.  Hofmeister  gives,  as  a  characteristic  of  melanins,  that  their 
elementary  molecular  composition  is  always  nearlv  in  the  proportions 
N    :   H    :   C  =  1    :  5    :  5. 

Gortner's  ^  studies  have  led  him  to  accept  the  general  principle  that 
melanin  is  formed  through  the  action  of  an  oxidase  on  an  oxidizable 
chromogen,  but  that  in  keratinous  structures  there  exist  at  least  two 
types  of  melanins,  one,  a  ''melano-protein,"  soluble  in  dilute  acids 
and  existing  dissolved  in  the  keratins;  the  other,  insoluble  in  dilute 
acids,  exists  as  pigment  granules  and  is  of  unknown  nature.  Piettre  '^ 
believes  that  melanin  from  sarcoma  of  the  horse  consists  of  a  protein 
united  to  a  pigment.  Those  whose  studies  of  melanin  formation  have 
been  made  with  the  microscope,  state  that  the  nucleus  is  active  in  the 
process,"  and  some  find  the  melanin  so  closely  related  to  the  lipoids 
that  they  consider  it  a  lipochrome.^ 

A  particularly  prominent  constituent  of  some  melanins  is  sulphur, 
which  has  been  found  in  as  high  proportions  as  10  per  cent,  in  mel- 
anin from  sarcomas,  and  even  12  per  cent,  in  sepia  from  the  squid ; 
in  melanin  from  hair  the  sulphur  is  usually  about  2-4  per  cent.;  but 
in  choroid  melanin,  and  in  some  other  forms,  sulphur  seems  to  be  ab- 
sent. The  proportions  of  sulphur  obtained  from  the  same  specimen 
purified   by   different  methods   show   wide   variations,   and  hence   v. 

4  Spioprlcr,  TTofmoistor's  Beitr.,  1007    (10),  253. 

•'".  Biofhom.  Bulletin.  liHl    (1),  207:  rr'suni^. 

oConipt.  Bend.  Acad.  Sci.,  1011  (I'l:^),  782;  also  sec  Ri'priiil  frnm  1st  Intor- 
nat.  ronfr.  Compar.  Pathol.,  Paris,  1912. 

T  StanVi,  Vcrli.  Df'iit.  Path.  Ges.,  1007  (11),  130;  Schultz,  Jour.  Mod.  Rca.. 
1912    (20),  05. 

sDvaon,  Jour.  Path,  and  Bact.,  1011  (15),  298;  KriMbieii.  Wien.  klin.  Woi'h., 
ion  "(24),  117. 


COMPOSITIOX  OF  MELAMX  469 

Fiirth  consiilers  that  iioitlier  the  .suli)hur  nor  the  iron  are  indispensa- 
ble constituents  of  the  melanin.  Probably  the  melanin  molecule  con- 
tains at(mi-coinploxes  that  have  a  tendency  to  bind  certain  sulphur 
and  iron  comi)()Uiids  (e.  g.,  cystine  or  hematin  derivatives). 

There  is  much  reason  to  believe  that  the  melanin  is  derived  from 
certain  p:roups  of  the  protein  molecule  that  seem  readily  to  form  col- 
ored compounds.  The  aromatic  compounds  of  the  protein  inolecule, 
such  as  tyrosine,  ]i]ienylalaiiine,  and  tryptophane,  readily  condense 
with  elimination  of  water  and  absorption  of  oxygen,  to  produce  dark- 
colored  substances.  When  proteins  are  heated  in  strong  hydrochloric 
acid,  we  obtain  a  dark-brown  material,  which  closely  resembles  the 
melanins  both  in  elementary  composition  and  in  general  properties, 
so  that  it  is  referred  to  as  "artificial  melanin"  or  "melanoid  sub- 
stance." These  substances,  like  the  natural  melanins,  when  decom- 
posed by  fusing  with  caustic  potash,  yield  skatole,  indole,  and  pyrrole 
derivatives,  which  are  undoubtedly  derived  from  the  tyrosine  and 
tryptopliane  of  the  protein  molecule.  Therefore,  it  seems  probable 
that  both  the  melanoid  substances  and  the  true  melanins  are  formed 
from  the  chromogen  groups  of  the  protein  molecule  through  processes 
of  condensation,  elimination  of  water,  and  the  taking  up  of  oxygen. ^^ 

Tyrosinase. — In  the  sepia  sacs  of  the  cuttle-fish,  in  meal-worms 
which  form  a  melanin-like  pigment,  and  in  plants  that  produce  the 
black  Japanese  lacquer,  have  been  found  oxidizing  enzymes  that  have 
the  property  of  producing  black  pigment  by  their  action  upon  tjTO- 
sine  and  other  aromatic  compounds.  Neuberg  °  found  that  extracts 
of  a  melanosarcoma  of  the  adrenal  could  produce  pigment  from 
epinephrin  and  ^-oxyphenylethylamine,  but  not  from  tyrosine.  The 
ink  sacs  of  the  squid  contain  an  enzyme  forming  a  pigment  from 
epinephrin,  apparently  through  oxidation  and  condensation.  These 
enz^'mes  may,  therefore,  possibly  be  responsible  for  the  production 
of  melanin  in  animal  tissues,  by  causing  oxidative  changes  in  the 
chromogen  groups  of  the  protein  molecule  that  are  liberated  by  auto- 
lysis (see  "Tyrosinase"  p.  73).  v.  Fiirth  urges  strongly  the  view 
that  both  normal  and  pathological  melanin  formation  depend  upon 
the  action  of  the  tyrosinase  or  allied  enzymes  in  conjunction  with 
autolytic  enzjones;  the  latter  split  free  the  chromogen  groups  of  the 
protein  molecule,  which  are  then  oxidized  by  the  tyrosinase,  undergo 
condensation,  and  take  up  sulphur-  and  iron-holding  groups  and  also 
other  organic  compounds,  the  entire  complex  forming  the  melanin. 

Properties  of  Melanin. — Wlien  isolated  in  a  pure  condition, 
melanin  is  a  dark-brown  substance  of  amorphous  structure,  no  mat- 
ter how  black  the  material  from  which  it  is  derived  may  be.^"     It  is 

SiiSee  Herzmark  and  von  Fiirtli,  Biochcm.  Zeit..  lill."?    (40),  130. 

9  Zoit.  f.  Krebsforsch..  IflO!)    (S),  195. 

10  Spiegler,  (Hofmeister's  Beitr.,  1903  (4).  40)  claims  to  have  isolated  from 
white  wool  a  white  chromogen,  closely  related  to  melanin  chemically,  but  Gortner 


470  PATHOLOGICAL    PIGMENTATION 

quite  insoluble  in  all  ordinary  reagents  except  alkalies,  in  which  some 
melanins  dissolve  easily,  and  some  with  difficulty.  Strong  boiling 
hydrochloric  acid  scarcely  affects  non-i)rotein  melanins.  By  the 
action  of  sunlight  or  oxidizing  agents  on  melanin-containing  sections 
the  pigment  can  be  bleached  out.  The  chief  decomposition-products 
formed  on  fusing  with  alkalies  are  indole,  skatole,  and  "melanic 
acid";  no  cystine,  leucine,  tyrosine,  or  other  amino-acids  can  be  iso- 
lated. ]\rost  authors,  therefore,  consider  the  melanins  as  heterocyclic 
compounds  standing  in  some  relation  to  the  indole  nucleus. 

If  melanin  is  injected  subcutaneously  into  animals  (rabbits  and 
guinea-pigs),  there  appears  in  the  urine  a  substance  which  turns  dark 
brown  after  the  urine  has  stood  for  some  time  (Kobert,  Helman). 
The  pigment  is  apparently  reduced,  particularly  by  the  liver,  to  a 
colorless  melanogen,  which  is  eliminated  in  the  urine.  The  same 
process  occurs  when  melanin  is  produced  in  excess  and  enters  the 
blood,  as  in  the  case  of  melanosarcoma,  a  colorless  melanogen  being 
formed  which  is  excreted  in  the  urine,  constituting  "melanuria." 
Occasionally  the  urine  is  dark  when  first  passed,  because  of  the  pres- 
ence of  melanin,  but  usually  it  must  be  subjected  to  oxidizing  agen- 
cies (bromine  water,  nitric  acid,  hypochlorites,  etc.),  or  exposed  to 
air  to  bring  out  the  brown  color.  Helman  ^^  says  that  true  melano- 
gen may  be  considered  to'  be  present  in  urine :  ( 1 )  If  the  careful 
addition  of  ferric  chloride  causes  the  development  of  a  black  precipi- 
tate. (2)  If  this  precipitate  dissolves  in  sodium  carbonate,  forming 
a  black  solution.  (3)  If  from  this  solution  mineral  acids  precipitate 
a  black  or  brownish-black  powder.  All  three  reactions  must  be 
obtained,  for  substances  other  than  melanin  may  give  the  first 
two. 

The  coloring  power  of  melanin  is  very  great,  for  urine  containing 
but  0.1  per  cent,  of  melanin  has  the  color  of  dark  beer  (Hensen  and 
Nfilke),  and  the  entire  skin  of  a  negro  contains  only  about  1  gram 
of  melanin  (Abel  and  Davis). ^-  Excessive  quantities  of  melanin  ma.y 
be  in  part  deposited  in  the  lymph-glands  and  skin,  causing  diffuse 
pigmentation ;  it  may  be  deposited  in  the  endothelium  lining  the 
blood-vessels,  hi  a  pigmented  colon  Al)derhalden  ^-''  found  melanin- 
like substances  which  seemed  to  be  derived  from  tryptophane.  Nik- 
las,'-''  however,  Ijelieves  t3a'osinase  activity  to  be  responsible  for  this 
type  of  intestinal  melanosis.  Kobert  injected  melanin  into  albino 
ra])l)its,  but  did  not  succeed  in  getting  any  dejiosition  in  the  choroid 

(Amor.  Xatiiralist,  1010  (44),  407)  boliovos  this  to  bo  a  docDmiKisif  imi  ])ro(liiot 
of  koratin,  iinrolalod  to  incdanin. 

iiCViit.  f.  inn.  Mod.,  1002  (2:5),  1017:  Aroh.  iiiloniat.  riianiial<odviiaiii..  100.3 
(12),  271. 

12  Jour.  Faj).  Mod.,  IHOG    (1),  .Sfil. 

i2aZoit.   phvsiol.   Choni..   101.3    (S.5).   02. 

i2b]\liinoli.  inod.  W(m-Ii..  1014  (61),  13:52.  8oo  also  llattori,  :\litt.  Mod.  Cosollscli. 
Tokio,  1010    (.30),  No.  6. 


MEI.AyorlC  TUMOItH  ■*'' 

„..  .kin      Ilclman  found  »onic  evidence  of  toxicity  when  lar^e  doses  of 
.ehn  in  d"    1  e      n  sodium  earbonate  are  injected  njto  an.mals.  but 

an,ns.     Iron  .s  £  •'^     "t  .Y  1™  ,an,ixture  of  blood-pipnent 

Crtr^nrol'c^a'troSanin^ba^sJroducedbythe^um^^^ 

comatou   lner.300grams^  states  that  the  melanin  may  con- 

'h,'  fte"7  3  per  c/nt  by  vve  ght  of  the  fresh  substance  of  some  melano- 
:ar:omIs    Tccorlfto  Utbarsch  and  to  Helman,  melauotrc  tumors 

'•^t';ritefZ:;Neuber.  found  that  a  "elastic  -coma;^^^ 

,a^eWense.»tlon^^ 

trvDtophane  was  tea  to  a  pdniiu.  v.  ^  \^^A\-  tn 

melamiria  He  therefore  concludes  that  tlie  power  of  the  bod>  to 
r.tr  the  Dvrrole  rin-  is  reduced,  and  instead  it  undergoes  reduc- 
tTLZ^^n,  ^ion  with  sulphuric  ^id  to  W  ^^^al 
s„lnhate  of  methvlpvrrolidlne-hydroxy-earbonic  acid  (CH,-b,li.lN..U4 ) . 
XbdJhalto-'  also  found  a  relation  to  tryptophane,  for  m  the  urme 
^f  atianuric  was  present  a  substance  rich  in  tr,-p.opl.ane ;  and 


13  \roh    internat.  Plmnnakodynam.,  1003    (1-). 

14  Jour.  Med.  Res.,   V.m    (16),   117. 
isVirchow's  Arch..   1000    (108).  62. 
16-Biochem.  Zeit.,  1010   (28).  ISl 
iTZeit.  physiol.  Chem.,  1912   (/8),  159. 


472  PATHOLOGICAL    PIGMEXTATION 

Priinavera  '^  found  the  iiriiie  in  a  case  of  melanosarcoma  contain- 
ing free  tyrosine,  fluctuating  in  amount  with  the  pigment. 

Addison's  disease  is  associated  with  the  deposition  of  a  pigment 
in  the  skin  that  is  generally  considered  to  be  a  melanin,  differing 
from  that  produced  normally  in  the  skin  only  in  quantity'  and  not  in 
origin  or  composition.^''  No  satisfactory  explanation  of  the  relation 
of  the  adrenal  to  this  pigmentation  seems  yet  to  have  been  made,  al- 
though it  is  natural  to  assume  that  when  the  function  of  the  adrenal 
is  destroyed,  substances  accumulate  in  the  blood  that  have  a  stimu- 
lating effect  on  the  pigment-forming  cells.  Abnormal  protein  catab- 
olism,  with  excessive  accumulation  of  the  chromogenic  constituents  of 
the  protein  molecule,  has  been  suggested,  as  also  have  alterations  in 
the  influence  of  the  sympathetic  nervous  system  upon  the  chromo- 
phore  cells,  for  nerve  lesions  (e.  g.,  neurofibroma)  often  are  accom- 
panied by  pathological  pigmentation  of  the  skin.-" 

It  is  significant  that  the  active  constituent  of  the  adrenal  medulla, 
the  epinephrin,  is  an  aromatic  derivative  closely  related  to  tyrosine, 
since  the  production  of  pigment  by  the  action  of  oxidizing  enzymes 
upon  such  substances  is  well  known.  Furthermore,  Neuberg  has  de- 
scribed a  melanotic  adrenal  tumor  which  produced  pigment  by  oxi- 
dizing epinephrin.  On  this  basis  the  pigmentation  of  Addison's  dis- 
ease would  seem  to  be  the  result  of  an  abnormal  accumulation  or  dis- 
tribution of  aromatic  compounds,  because  of  their  failure  to  be  con- 
verted into  epinephrin.  In  support  of  this  hypothesis  is  the  obser- 
vation of  ^leirowsky  that  the  human  skin  contains  an  enzyme  capable 
of  oxidizing  e])inephrin  to  a  pigment,  and  that  pieces  of  skin  kept 
warm  will  develop  a  postmortem  pigmentation,  and  this  is  supported 
by  Konegstein  -^  who  found  that  the  pigmentation  was  greater  in 
animals  deprived  of  their  adrenals  or  given  injections  of  epineph- 
rin. 

As  exact  chemical  studies  of  the  pigment  in  Addison's  disease  have 
not  been  made,  however,  we  have  no  positive  proof  that  it  is  a  mela- 
nin, hence  any  speculation  as  to  the  cause  of  its  formation  is  prema- 
ture. Carbone --  claims  to  have  isolated  from  the  urine  in  Addison's 
disease  a  pi^iment  that  contains  much  suljiliur,  and  which  he  considers 
similar  to  or  identical  with  the  melanogen  of  melanuria.  A  similar 
observation  is  reported  by  Eiselt.--'  v.  Kahlden,-^  however,  has  ob- 
served crystals  resembling  hematoidiii  in  the  pigmented  tissues. 

isfiiorn.  Int.  Scicnze  Med.,  1908   (29),  978. 

19  Concerning  histogenesis  of  the  pigment  see  Pfuiringcr,  Cent.  f.  Path.,  1900 
(11),  1. 

20  See  r68iim^  hv  Schmidt,  Ergeb.  der  Pathol.,  1896  (Bd.  ,3.  Abt.  1),  551. 
-■1  Wicn.  l<lin.  Woch.,  1910   (2.3),  GIG. 

22  (Jioino  H.  Acad.  nied.  di  Torino,  1S9G. 

2't  Zeit.  kiln.  Med..  1910  ( G9 ) .  393 ;  full  discussion  on  the  pigment  of  Addison's 
disease. 

24  Virchow's  Arcli.,    1SS8    (114),  Go. 


OCHRONOSIS  473 

Ochronosis  -^  is  a  condition  characterized  by  a  black  pigmentation 
of  the  cartilages,  first  described  by  Virchow  in  1866.  In  1904  Osier  ^'' 
reported  two  eases,  and  fonnd  bnt  .seven  others  in  the  literature  to 
that  time.  The  origin  and  nature  of  tliis  pigment  remains  still  un- 
decided. Virchow  suspected  that  the  condition  was  due  to  a  permea- 
tion of  cartilage  by  hematin  derivatives,  but  Hansemann,  finding  a 
case  associated  with  melanuria,  considered  that  the  pigment  is  prob- 
ably of  metabolic  origin.  Ilecker  and  AYolf  studied  the  urine  of  a 
similai'  case,  and  concluded  that  the  pigment  must  be  melanin.  Al- 
brecht,-^  however,  suggested  a  relation  of  ochronosis  to  alkaptonuria, 
having  found  homogentisic  acid  in  the  urine  of  a  case  reported  by 
him  (see  "Alkaptonuria").  Osier's  two  patients  were  brothers  with 
alkaptonuria,  the  evidence  of  ochronosis  consisting  of  discoloration  of 
the  cartilages  of  the  ears.  Langstein  -**  has  examined  a  specimen  of 
urine  preserved  from  Hansemann 's  case,  and  found  no  evidence  of 
alkaptonuria.'" 

Pick  ^'^  summarizes  the  results  of  his  study  of  his  case  and  of  the 
literature,  as  follows :  Ochronosis  is  a  definite  form  of  melanotic  pig- 
mentation, the  pigment  of  ochronosis  being  in  most  of  the  cases  very 
closely  related  to  melanin.  The  pigment,  or  its  chromogen,  circulat- 
ing freely  in  the  blood,  is  imbibed  not»only  by  cartilage,  but  also  by 
loose  connective  tissue,  voluntary  and  involuntary  muscle-cells,  and 
epithelial  cells,  without  any  decrease  in  vitality  of  these  cells  being 
observable ;  however,  degenerated  tissues  show  the  greatest  amount  of 
pigmentation.  The  diffuse  pigment  can  become  granular  after  a  time ; 
it  is  iron-free,  but  under  certain  circumstances  may  contain  fat. 
Thi^  melanin  arises  from  the  aromatic  nucleus  of  the  protein  mole- 
cule (tyrosine,  phenylalanine),  and  the  related  hydroxylized  products, 
binder  the  influence  of  tyrosinase.  In  some  cases  the  constant  ab- 
sorption of  minute  quantities  of  phenol  from  surgical  dressings  seems 
to  have  been  the  cause  of  the  condition.  Besides  this  formation  of 
pigment  from  such  "exogenous"  aromatic  substances,  however,  it  is 
probable  that  in  alkaptonuria  the  "endogenous"  aromatic  substance 
(homogentisic  acid)  present  may  be  converted  into  pigment  by  the 
tyrosinase.  In  many  of  the  cases  of  ochronosis  the  pigment  or  a 
precursor  may  be  excreted  in  the  urine,  which  then  undergoes  spon- 
taneous darkening  when  exposed  to  the  air.  The  kidneys  may  also 
become  pigmented  and  granular  masses  of  pigment  may  be  present 
in  the  renal  tubules.   . 

25  See  Adler,  Zeit.  f.  Krebsforsch.,  1911  (11),  1:  Poulsen,  Ziegler's  Beitr.,  1010 
(48).  346. 

26  Lancet.  1004    (i),  10    (literature). 

27  Zeit.  f.  Heilk.,  Path.  AM.,  1002   (2.3),  366. 
2s  Hofmeister's  Beitr.,   1003    (4),  145. 

20  Also  see  Langstein.  Berl.  klin.  Woch..   1006    (43),  507. 
30Berl.  klin.  Wochenschr.,  1906    (43),  478. 


474  PATHOLOGICAL    PIGMENTATION 

Poulsen  "  states  that  of  the  32  known  eases  of  oehronosis  (in  1911) 
in  17  there  was  alkaptonuria,  in  8  carbolic  acid  dressings  had  been 
used  for  long  periods,  and  in  the  remaining-  7  cases  the  cause  was  not 
determined.  These  facts  are  conclusive  evidence  of  the  origin  of 
ochronotic  pigment  from  aromatic  radicals,  whether  these  radicals 
are  converted  into  true  melanin  or  not.  The  localization  of  the  pig- 
ment is  explained  by  the  demonstration  by  Gross  and  Allard,^-  that 
cartilage  has  a  greater  affinity  than  other  tissues  for  homogentisic 
acid.  There  are,  however,  numerous  cases  of  alkaptonuria  without 
ochronosis.  The  ochronosis  described  in  lower  animals  is  not  the 
same  as  human  ochronosis,  affecting  the  bones  rather  than  the  carti- 
lages (Poulsen), 2^  and  being  more  properly  designated  by  the  name 
osteohemachromatosis  (Schmey).^* 

Malarial  pigmentation,  according  to  Ewing,^^  may  have  any  one 
of  the  following  origins : 

(1)  Pigment  elaborated  by  the  intracellular  parasite.  (2)  Hem- 
atoidin  derived  from  the  remnants  of  infected  red  cells.  (3)  Hem- 
atoidin  or  altered  hemoglobin  deposited  in  granular  or  crystalline 
form  from  red  cells  dissolved  in  the  plasma.  (4)  Bilirubin  or  uro- 
bilin granules  or  crystals. 

Of  these,  the  pigment  formed  by  the  parasites  has  been  considered 
by  many  as  a  true  melanin,  but  this  cannot  be  considered  as  estab- 
lished, especially  as  Ewing  finds  it  to  have  the  same  relation  to 
solvents  as  do  the  blood-pigments.  Carbone  and  Brown  ^^  consider 
the  malarial  pigment  to  originate  from  hematin,  with  which  it  agrees 
in  solubility,  spectroscopic  properties,  and  in  containing  iron. 

LIPOCHROME 

In  normal  plant  and  animal  tissues  occur  pigments  that  are  either 
fats  or  compounds  of  fat,  or  substances  highly  soluble  in  fats.  In 
animals  they  occur  normally  in  the  corpus  luteum,  in  the  epithelium 
of  the  seminal  vesicles,  testicles,  and  epididymis ;  in  ganglion-cells, 
especially  in  the  sympathetic  nervous  tissue ;  in  the  Kupffer  cells  of 
the  liver;  and  in  fat  tissue.  Pathologically,  such  pigments  are  found 
particularly  in  the  muscle-cells  in  brown  atro])hy  of  the  heart,  and 
less  abundantly  in  the  epithelium  of  atrophied  livers  and  kidneys 
(Lubarsch  ^^  and  Sehrt  ^^).  All  are  characterized  by  staining  by  such 
fat  stains  as  Sudan  III  and  scarlet  R,  and  usually,  but  not  constantly, 

3iMuneh.  mod.  Wodi..   1012    (50).  .'304. 

32  Arch.  e\p.  Path.  u.  Pliarm.,   1008    (.^O),  .'3S4. 

33  See  Tn<rior,  Ziofjlor's  Boitr..  1011    (r>l).  100. 

34  Frankfurtor  Zoit.  Patliol.,  lOl.S  (12),  21S;  also  Toutsclilaoiuler.  Virdiow's 
Arch.,  1014    (217).  .-^O.^. 

35  .Tour.  Exp.  l\To(l..  1002    (fi),  110. 

30  Jour.  Exppr.  Med.,   1011    (1.3).  200. 
37  Cent.  f.  Pathol.,   1002    (13).  SSI. 

3SVirchow's  Ardi.,  1004  (177),  24S.  S(>o  also  Afavor  et  ah.  Jmir.  plnsiol.  ct 
path.  ji<-"n.,  1014    (16),  581. 


L/I'OCHROME  475 

by  osiiiic  aeitl ;  they  are  dissolved  by  the  usual  fat  solvents.  It  is 
questionable  if  all  pigments  that  stain  for  fat  should  be  considered  as 
true  lipochromos,  however,  for  their  other  reactions  are  variable; 
and  Rorst  would  distin<;-uisli  these  path()lo<i-i('al  pigments  from  the 
true  lipochromes  by  calling  them  lipofuscins,  including  under  this 
term  the  brown  "waste  pigments,"  which  Hueck  believes  to  be 
formed  from  disintegrated  lipoids  or  fatty  acids.  Many  pigmentary 
substances  are  probably  soluble  in  fats,  and  in  this  way  the  lipofus- 
cins are  formed.'''"'  Ty})ical  plant  lipochromes,  inchuling  the  pig- 
ments of  Staphylococcus  pyogenes  aureus  and  citreus,  are  colored  blue 
by  concentrated  sulphuric  acid  with  formation  of  small  blue  crystals 
of  lipocyanin.  With  iodin-potassium-iodide  solution  they  are  col- 
ored green.  Lipochrome  of  frog-fat  stains  blue  with  the  iodin-potas- 
sium-iodide solution  (Neumann)  ;  •''•*  lipochrome  of  the  corpus  luteum 
(called  lutein)  occasionally  gives  a  faint  blue  with  sulphuric  acid  or 
Lugol's  solution  (Sehrt)  ;  but  the  fat-holding  pigments  of  the  other 
tissues  mentioned  above  do  not  give  either  of  these  reactions.  Pos- 
sibly these  last  are  not  true  lipochromes,  therefore,  but  rather  pig- 
ments chemically  or  physically  combined  with  fat.  Cotte  *°  believes 
that  the  true  lipochromes  of  plants  and  animals  have  a  cholesterol 
base,  but  the  presence  of  glycerol  in  plant  and  bacterial  lipochromes 
can  be  demonstrated  by  the  acrolein  test — possibly,  therefore,  both 
cholesterol  and  neutral  fats  are  present.  iMelanins  and  pigments  de- 
rived from  hemoglobin  do  not  stain  with  Sudan  III  and  are  not  soluble 
in  ether,  etc.,  and  hence  can  be  readily  distinguished  from  the  fatty 
pigments.  It  has  been  shown  by  Escher  **'"*  that  the  pigment  of  the 
corpus  luteum  is  identical  with  the  carotin  of  carrots.  Apparent!}' 
carotin  and  xanthophyll  (a  crystalline  pigment  from  green  plants)**"' 
are  the  chief  pigments  of  milk  fats,  e^^  yolk,  and  probably  of  body 
fats.^^  In  the  bod}^  lipins  these  pigments  accumulate  throughout 
life  because  of  their  great  solubility  in  liiiins,  which  explains  the 
high  color  of  the  fats  of  old  persons.  Carotin  seems  to  be  almost  or 
quite  devoid  of  toxicity.*^''  In  the  renal  epithelium  is  found  a  pig- 
ment resembling  the  lipofuscins,  increasing  with  age  and  not  related 
to  the  urinary  pigments.*^*^  The  pigment  of  nerve  cells  resembles 
that  produced  during  autolysis  in  ganglia. •*^'= 

The  pigment  that   causes  the  peculiar  green   color  characteristic 

asa  Ciaccio  (Biochem.  Zeit..  1015  (fiO),  .31.3)  agrees  with  Hueck,  and  finds  it 
possible  to  distinguisli  between  pigments  from  phospliatids,  whieli  stain  poorly 
with  Sudan  III,  and  tliose  from  free  fattv  acids  which  stain  deeply  with  tiiis  dve. 

39Virchow's  Arch..    1002    (170),  363. 

•loCompt.  Rend.  Soc.   Biol..   1003    (5.1).  S12. 

4oaZeit.  phvsiol.  Chem..  1013   (S3),  lOS. 

40b  Concerning  plant  pigments  see  review  bv  West  and  Horowitz,  Biocliem. 
Bullet.,  1915    (4),   1.51  and  101. 

41  See  articles  by   Palmer  and  Eckles,  Jour.   Biol.  Chem.   1014,  Vol.   17   et  seq. 

4ia  Wells  and  Hedenburg,  .lour.  Biol.  Chem.,  1016    (27),  213. 

41b  Schreycr,  Frankf.  Zeit.  Pathol.,  1014   (1.5).  333. 

41C  Marinesco,  C.  R.  Soc.  Biol.,  1013   (72),  838. 


476  PATHOLOGICAL    I'KlMEXTATIOy 

of  certain  nialignant  g-rowtlis,  chloroma,^-  was  considered  by  Chiari, 
Huber  and  others  as^a  fatty  substance  related  to  or  identical  with  the 
lipochromes.  It  commonly  fades  on  exposure  to  air,  and  also  when 
in  the  usual  preservative  fluids,  to  which  it  does  not  impart  its  color. 
The  color  may  be  brought  back  after  formaldehyde  preservation  by 
H2O2  or  by  weak  alkalies  (Burgess).^-  Ottenberg  "'^  has  suggested 
that  the  green  color  may  be  due  to  eosinophiles  which  abound  in 
ehloromas,  since  in  fresh  preparations  eosinophile  granules  have  a 
faint  greenish  tinge.  It  contains  no  iron,  is  soluble  in  absolute  alco- 
hol and  in  ether,  and  is  usually,  but  not  always  (v.  Recklinghausen), 
stained  black  with  osmic  acid."'*  Treadgold  states  that  as  the  green 
color  is  not  present  from  the  beginning  it  Avould  seem  that  cellular 
degeneration  mu.st  play  a  part.  Possibly  a  degeneration  of  the  gran- 
ules of  the  mj'eloeytes  and  myeloblasts,  aided  by  the  products  of  hemo- 
globin disintegration,  is  responsible.**^ 

Chromophile  cells  may  be  considered  in  this  connection.  Kolin  45  has  described 
certain  cells  witli  a  decided  affinity  for  chromic  acid  and  its  salts,  found  abun- 
dantly in  the  sympathetic  nervous  system,  in  the  carotid  o-land.  and  in  the  medulla 
of  the  adrenal.  Tliey  are  also  present  in  tumors  derived  from  tiiese  organs. 
Extracts  from  sucli  organs  have  a  marked  effect  in  raising  blood  pressure,  and, 
according  to  Wiesel,-»6  they  are  greatly  involved  in  Addison's  disease.  The  nature 
of  the  chromophile  substance  is  unknown,  but  it  can  only  be  fixed  by  chromic 
acid  or  chromates;  cells  liardened  by  other  means  show  merely  spaces  in  the 
places  occupied  by  this  substance.  It  is  generally  believed  to  be  the  same  as  the 
epinephrin.  but  it  does  not  always  parallel  in  amount  the  quantity  of  epinephrin 
as  determined  chemically. 

BLOOD  PIGMENTS  47 

Red  corpuscles  behave  much  as  do  other  non-nucleated  fragments 
of  cells,  undergoing  disintegration  rapidly  and  constantly  when  un- 
der normal  conditions,  as  well  as  when  subjected  to  various  harmful 
influences  (see  "Hemolj^sis"),  or  when  outside  of  the  vessels  in  ex- 
travasations of  blood.  The  processes  and  products  of  their  disinte- 
gration are,  therefore,  much  the  same  whether  occurring  under  normal 
or  pathological  conditions.  The  hemoglobin  molecule  is  large  and 
complex,  and  from  it  are  derived  many  substances  of  the  nature  of 
pigments;  indeed,  hemoglobin  itself  may  appear  free  as  a  pigment. 

Hemoglobin  is  a  compound  protein,  consisting  of  a  protein  group 
(glohiii)  and  a  coloring-matter  {hematin  or  hemochromogen) .  The 
protein  globin  is  of  a  basic  nature,  and  seems  allied  to  the  histons. 

42  Literature  bv  Dock.  Amer.  Jour.  Med.  Sci..  1S9.3  (100).  152;  and  Dock  and 
Warthin,  Med.  News,  1004  (85),  971;  Burgess,  Jour.  IMcd.  Res.,  1912   (27),  13.3. 

43  Amer.  Jour.  Med.  Sci.,  1909    (138),  505. 

44  Tlie  pigineiit  of  xnnthclntima  multiplex  seems  to  be  a  fattv  substance  (  Poens- 
gen.)      Virchow's  Arch.,  1883    (91),  354. 

44a  Quart.  Jour.  Med.,  1908  (1),  239;  Weber,  Proc.  Hov.  Soc.  :\red.,  Clin.  Med. 
Sec,  1916   (0),  7. 

45Prag.  med.  Woch.,  1902   (27),  325. 

46Zcit.  f.  Heilk.,  Path.  Abt.,  1903   (24),  257. 

47  Literature  bv  Schmidt,  Krgebnisso  dor  Pathol.,  1894  (L),  101;  and  1896 
(III,),  542;   Sdiulz,  Ergebnisse  dor  Physiol.,  1902    (I,),  505. 


BLOOD  PIGMENTS 


477 


The  liematiu  is,  tlierofore,  presumably  acid,  and  tlie  coiiipound  pro- 
tein, hemoglobin,  is  somewhat  like  the  nucleoproteins  in  nature.  Hem- 
oglobin ordinarily  does  not  crystallize  readily,  especially  the  hemo- 
globin of  man,  and  it  is  doubtful  if  it  ever  does  so  in  the  living 
tissues,  although  possibly  this  may  occur  in  the  center  of  large  hema- 
tomas. In  bodies  that  have  undergone  postmortem  decomposition, 
and  occasionally  in  specimens  kept  for  microscopic  purposes,  irregu- 
lar orange-yellow  crystalline  masses  of  hemoglobin  may  be  found. 
Tliis  occurs  particularly  if  the  blood  has  been  acted  upon  by  hemo- 
lytic agents  or  has  undergone  i)utrefactive  changes,  and  then  is 
hardened  in  alcohol.  The  crystals  are  either  oxyhemoglobin,  or  more 
often  an  isomeric  or  polymeric  modification,  parahemoglohin  (Nen- 
cki).  Hemoglobin  also  enters  cells  unchanged,  imparting  a  dififuse 
yellowish  color,  and  apparently  it  is  non-toxic.'*"'^  If  present  in  the 
blood  in  large  enough  amounts  it  is  excreted  unchanged  in  the  urine, 
but  at  least  one-sixtieth  of  the  total  number  of  red  corpuscles  must 
be  in  solution  at  one  time  to  produce  hemoglobinuria;  in  man  at 
least  17  e.c  of  laked  corpuscles  must  be  injected  to  accomplish  this.'*''' 

Addis  *'"  has  developed  the  following  conception  of  the  metabolism 
of  hemoglobin.  Free  hemoglobin,  liberated,  especially  by  the  phago- 
cytes of  the  spleen,  is  taken  up  by  the  other  phagocytes,  notably  the 
Kupffer  cells  of  the  liver,  which  pass  it  on  to  the  liver  cells.  The 
])igment  moiety,  hemochromogen,  is  separated  from  the  globin,  and 
converted  through  removal  of  its  iron  into  hiliruhin.  The  bilirubin 
excreted  into  the  intestine  is  there  reduced  to  urobilinogen,  w'hich  is 
in  part  reabsorbed  and  polymerized  into  uroMlin,  which  in  turn-  is 
possibly  polymerized  into  a  larger  complex.  In  the  liver  this  uro- 
bilin complex  has  restored  to  its  pyrrol  nuclei  the  original  side  chains, 
and  then  is  used  to  form  new  hemoglobin  molecules.  This  hypoth- 
esis is  merely  tentative,  but  it  affords  a  useful  "working  hypoth- 
esis" for  the  consideration  of  many  phases  of  pigment  metabolism. 

In  the  decomposition  of  hemoglobin  the  first  step  is  the  splitting 
of  the  globin  (which  does  not  form  pigments)  from  the  hematin,  from 
which  many  pigments  may  be  derived. 

Hematin. — The  formula  given  for  this  substance  by  Neneki,. 
C.^nHooN^Fe04,  has  been  generally  accepted,  although  it  is  not  cer- 
tain that  the  hematin  of  all  animals  is  the  same.  It  is  found  fre- 
quently as  an  amorphous,  dark-brown  or  bluish-black  substance,  in 
large,  old  extravasations  of  blood,  but  seldom  in  small  hemorrhages. 
As  a  pathological  pigment,  however,  hematin  is  by  no  means  so  fre- 
quently found  as  its  derivatives.  Schumm  ^^  observed  a  patient  with 
chromium  poisoning  wdio  showed  for  several  days  abundant  hematin 
free  in  the  blood.     He  has  also  found  it  in  malaria,  pernicious  anemia 

4TaBarratt  and  Yorke,  Brit.  Med.  Jour.,  Jan.  .31,  1914. 

47bSellards  and  IMinot,  Jour.  Med.  Res.,  1916    (34),  4G9. 

4Tcj\rch.  Int.  Med..  1913    (1.5),  412. 

48Zeit.  phvsiol.  Chem.,  1912   (80),  1;   191.3    (87).  171;   1916    (97),  32. 


478  PArilOLOCIVAL  PIGMEXTATIOX 

and  generalh-  with  acute  toxic  heiiioh'sis,  including  patients  infected 
with  B.  c))iphi/scmatosus,  when  the  hematin  may  be  accompanied  by 
methemogk)biu  without  a  corresponding-  urinary  excretion  of  these  pig- 
ments. Brown  *^  found  that  solutions  of  hematin  cause  chills  and 
fever,  and  suggests  tliat  his  pigment  may  be  at  least  partially  respon- 
sible for  the  symptoms  of  malaria/""  Hematin  has  been  believed  to 
split  up  gradually  into  an  iron-free  pigment  {hematoidin)  and  an 
iron-containing  pigment  (hemosiderin).  This  change  may  be  repre- 
sented by  the  following  equation,  according  to  Nencki  and  Sieber :  ^'^ 

C3=H3,N,0,Fe  +  2H,0  =  2C,„H,,K.03  +  Fe. 
(homatin)  (liematoidin) 

However,  finding  that  the  pigment  in  the  malarial  spleen  is  hem- 
atin, Brown  ^^  suggests  that  hematin  cannot  well  be  an  intermediary 
])roduct  in  hemoglobin  disintegration,  since  this  malarial  pigment 
])ersists  a  very  long  time  in  the  tissues  without  change.  He  has  made 
other  observations  that  led  him  to  conclude  that  hematin  is  not  an 
intermediary  substance  between  hemoglobin  and  hemosiderin,  but  that 
when  once  formed  it  is  destroyed  very  slowly,  by  oxidation  rather 
than  hydrolysis.  Injected  into  rabbits  it  produces  vascular  lesions  in 
the  kidneys  ^^^  and  in  large  doses  causes  a  marked  fall  in  blood  pres- 

g^^pp  r,ib 

Hematoidin  may  be  found  in  old,  large  extravasations,  as  orange- 
colored  or  red  rhombic  plates,  first  described  bj'  Virchow\  Some- 
times, however,  hematoidin  occurs  in  the  form  of  yellowish  granular 
masses,  and  it  may  be  associated  with  lipoids;  it  is  also  found  in  crys- 
talline form  in  icterus  (Dunzelt).^-  It  seems  to  be  nearly  or  quite 
identical  with  the  bile-pigment,  bilirubin,  and  it  is  probably  the 
source  of  this  substance  under  nornuil  conditions.  When  formed  in 
excessive  amounts,  either  through  increased  destruction  of  corpuscles 
in  the  vessels  or  in  extravasations,  the  amount  of  bile-pigment  is  in- 
creased (see  "Icterus").  Possibly  some  of  the  hematoidin  becomes 
transformed  directly  into  urobilin,  and  is  tlien  eliminated  in  the 
urine. 

Hemosiderin  ■'^'  is  relatively  insoluble,  and,  therefore,  is  more 
slowly  removed  when  formed  in  hemorrhages,  and  more  abundantly 
deposited  in  the  tissues  when  formed  after  excessive  hemolysis.  In 
acute  hemolytic  anemia  a  third  of  the  total  iron  of  the  blood  may  be 

49  Jour.  Exppr.  Med..  1012    (1.5),  ;i80;    191.3    (18),  96. 

4»a  Disputed  by  Butterfiold  and  Bciipdict,  Prof.  Soc.  Exp.  Biol..  1914    (11),  80. 

no  Arch.  cxp.  Patli.  \\.  Pharm..  ISSS  (24),  440;  Pruf^scli  and  Yoshimoto,  Zeit. 
cxp.  Path..  1911    (8),  039. 

niJour.  Expor.  Mod.,   1911    (1.-?),   290;    1911    (14),  (;12. 

.-.laArdi.  Int.  M.d.,  1913    (12),  315. 

51b  Brown  and  T>o('V(>nliart,  Jour.  Exp.  ^Nlcd.,  1913   (IS),  107. 

52  Cent.  f.  Path..  1909    (20).  900. 

5.3SPO  Xeun.ann,  Vircliow's  Arch.,  1888  (111),  25;  1900  (101),  422;  1904 
(177),  401;  also  Arnold,  ihid.,  1900  (ICl),  284;  Leupold,  Beitr.  path.  Anat.,  1914 
(59),  501. 


jii.oon  I'KiM i:\TS  479 

deposited  in  the  liver,  spleen  and  kidneys  within  2-1  hours.''^''  In  in- 
farcts hemosiderin  soon  disappears  (Schmidt),'^*  presumably  because 
dissolved  by  the  acids  formed  during-  autolysis.  According-  to  Neu- 
mann, hemosiderin  is  produced  only  under  the  influence  of  living  cells 
and  in  the  presence  of  oxygen,  while  hematoidin  arises  independent  of 
cellular  activity ; '"''  but  ]^rown  ■'''  has  found  that  hemosiderin  can  be 
formed  during  autolysis  of  the  liver,  especially  when  air  is  present, 
and  thei-efore  pi'obably  by  an  oxidizing  enzyme.  He  suggests  that  in 
hemosiderin  the  pigment  is  still  hematoidin,  and  that  the  formation 
of  hemosiderin  takes  place  in  the  nuclei,  the  hemosiderin  being  made 
directly  from  hemoglobin  without  the  intervention  of  hematin.  It 
may  also  be  formed  from  the  iron-containing  protein  of  the  cells  dur- 
ing autolysis,  independent  of  hemoglobin."'"  jNIilner  ^*  considers  that, 
under  similar  conditions,  an  iron-containing  pigment  is  also  formed, 
which  differs  from  hemosiderin  in  having  the  iron  so  combined  that 
it  cannot  react  with  the  usual  reagents ;  this  pigment  may  later  change 
into  hemosiderin.  Up  to  the  present  time  we  do  not  know  the  chem- 
ical nature  of  hemosiderin,  nor  its  exact  fate  in  the  body,  but  it  is 
probabl}'  utilized  in  the  manufacture  of  new  hemoglobin,  for  it  is 
known  that  the  iron  liberated  when  hematin  is  broken  up  in  the  body 
under  experimental  conditions  is  deposited  and  not  eliminated  (Mor- 
ishima).'"'^ 

Unstained  hemosiderin  generally  appears  in  the  form  of  brown 
or  yellowish-brown  granules,  and  not  as  crystals.  After  a  time  it  is 
taken  up  and  deposited  to  a  large  extent  in  the  liver,  spleen,  bone- 
marrow,  and  kidney,  either  as  hemosiderin  or  possibly  as  some  other 
iron  compound  of  similar  nature.  From  these  sites  it  seems  to  be 
later  taken  up  to  be  utilized  in  the  manufacture  of  new  red  cor- 
puscles. 

All  told  the  average  human  body  contains  about  3.2  grams  of  iron, 
of  which  2.4  to  2.7  grams  is  in  the  blood.  According  to  Meyer  ""^ 
iron  is  present  in  the  body  in  three  forms :  1.  Not  demonstrable  by 
reagents  because  so  firmly  bound  (hemoglobin).  2.  Loosely  bound 
iron,  colored  by  (NIT4)oS  acting  for  a  long  time  (ferratin).  3.  Salt- 
like compounds  with  proteins,  and  inorganic  compounds,  reacting  at 
once  with  reagents.  Ferratin  is  the  iron  compound  in  the  liver,  con- 
taining 6  per  cent.  iron.  He  believes  that  probably  hemosiderin  is 
not  a  definite  substance,  but  merely  indicates  compounds  of  the  third 

33aMiiir  and  Dunn.  Jour.  Path,  and  Baot.,  1015    (10),  417. 

54  Verb.  Deut.  Path.  Cesell.,  lOOS    (12),  271. 

55  The  aocuninlation  of  iron  in  tho  liver  wliich  follows  poisoninijf  with  hemolytic 
agents,  is  not  prevented  or  diminished  bv  preliminary  removal  of  the  spleen 
(Meinertz,  Zeit.  exp.  Path.  u.  Ther.,  1906    (2),  602). 

56  Jour.  Exper.  Med.,  1910   (12),  623. 

5T  Sprunt  et  al.,  Jour.  Exp.  Med.,  1912    (16),  607. 
ssVirchow's  Arch.,  1903    (174),  475. 
59  Arch.  exp.  Path.  u.  Pharm.,  1898    (41),  291. 
eoErgeb.  der  Physiol,  1905   (5),  698;  literature. 


480  PATIIOLOaiCAL    PIGMEXTATIOX 

class.  Iron  pigments  may  be  transformed  from  one  class  to  another, 
e.  g.,  in  corpus  lutenm  scars,  whose  age  can  be  estimated,  class  three 
may  be  replaced  by  class  two.  We  may  have  in  the  sputum  and  lungs 
"  Herzf ehlerzellen "  that  either  do  or  do  not  stain  with  ferrocyanide. 
In  morbus  macidosus,  Kunkel  found  the  pigment  of  the  internal  or- 
gans to  be  pure  iron  oxide.  Hueck  also  hohls  that  hemosiderin  is  an 
inorganic  iron  compound,  loosely  bound  to  proteins  and  fats,  and 
that  it  never  forms  an  iron-free  pigment,  as  has  been  stated.  He  be- 
lieves that  there  is  very  little  iron  in  the  tissues  in  a  firm  union  like 
hemoglobin,  and  that  by  proper  technic  some  iron  can  be  stained  in 
every  organ  which  contains  iron  chemicallj'  demonstrable.  Ischida  '^^ 
believes  that  an  iron-containing  pigment  may  be  formed  in  striated 
muscles  from  the  iron  normally  there,  w^ithout  requiring  a  hematoge- 
nous origin. 

Hematoporphyrin."- — There  are  several  closely  related  pigments  de- 
rived from  hematin  that  are  appropriately  grouped  under  the  desig- 
nation of  porphyrins,  for  they  are  not  all  identical  with  the  pigments 
prepared  artificially  from  hematin  b}^  Nencki  and  called  by  him 
heniatoporphyrin  and  mesoporphyrin,  the  former  apparently  repre- 
senting a  reduction,  the  latter  an  oxidation  product. ''^  The  porphy- 
rins found  in  the  urine  and  feces  are  different  from  each  other  and 
from  those  prepared  by  Nencki.*'*  Physiologically,  these  pigments  are 
of  great  interest,  because  of  the  close  chemical  relation  they  have  been 
found  to  bear  to  chlorophyll,^'^  with  which  hemoglobin  is  so  closely  re- 
lated functionally.  It  is  also  interesting  to  consider  that  whereas  car- 
nivora  obtain  much  hemoglobin  in  their  food,  herbivora  obtain  much 
chlorophyll.  Pathologically,  porphyrin  is  of  interest  as  a  urinary 
pigment,  being  found  normally  in  the  urine  in  traces,  but  present  in 
considerable  quantities  in  many  diseases,'^*'  such  as  rheumatism,  tuber- 
culosis, various  liver  diseases,  and,  most  strikingly,  after  the  admin- 
istration of  sulphonal,  veronal  or  trional.  When  in  abundance  it 
may  color  the  urine  a  rich  Burgundy  red,  and  it  is  sometimes  accom- 
panied by  a  precursor,  vro-fuscin.  It  is  present  in  the  bones  of  animals 
showing  hemochromatosis  and  in  the  bones  of  persons  *""'  exhibiting  a 
congenital  form  of  "hematoporphyria, "  described  by  Giinther,  which 
is  accompanied  by  severe  skin  lesions  that  are  ascribed  to  the  action  of 
light  upon  the  skin  sensitized  by  the  hematoporphyrin.  Hausmann  •'^ 
and  others  have  studied  extensively  the  photosensitizing  action  ex- 

«i  Virehow's  Arch.,  1012   (210),  67. 

"s  Literature  and  full  review  bv  Giintlier,  Deut.  Arcli.  kiiu.  :\[ed.,  1012  (105), 
80;  and  by  Jesionek,  Erjreb.  inn.  Med.,   1013    (11),  .52"). 

«■■!  Fischer  and  Meyer-Bet/,  Zeit.  physiol.  Chem.,  1912   (82).  06. 

04  11.  Fischer,  Miiiich.  nied.  Woch..  1016  (63).  377;  Zeit.  phvsiol.  Chem.,  1016 
(97),  100  and  148;   Schumni,  ibid.,  1015    (06).   183. 

«5  For   literature  see  Aliderhalfh'ii,  "Lehrbucli   der   plivsiol.   C'lieniie."   1000. 

«<■■  See  Carnid,  .lour,  of    Phvsiol.,    1802    (1:5),   .'"jOS. 

O'iallegler  ct  a/.,  J)eut.  iiied.  Wodi.,   1013    (30),  842. 

07  Biochcm.  Zeit.,  1010    (3(1),  27(;;   1014    (67),  300. 


PSEUDOMELANOSIS  481 

Libited  by  liematoporpbyrin  and  other  porphyrins,  and  find  evidence 
suggesting  a  rehitionship  between  hematoporpliyria  and  "hydroa 
aestiva,"  and  other  conditions  in  which  the  skin  is  abnormally  sensi- 
tive to  light. 

After  injection  of  0.2  gm.  hematoporphyi-in  into  his  own  veins, 
Mej'^er-Betz  '^'^^  found  himself  so  sensitized  to  light  that  exposure  to 
the  sun  caused  severe  skin  reactions  during  a  period  of  weeks,  and 
exposure  to  the  Finsen  light  produced  severe  ulceration ;  but  little 
hematojiorphyrin  escaped  in  the  urine.  Many  other  products  of 
blood  destruction  tested  on  animals  were  without  sensitizing  effects. 
Meth^-lation  of  the  p^'rrol  groups  only  partially  removes  the  activity 
of  hematoporphyrin.  Porphyrin  obtained  from  urine  and  feces  by 
Fischer  also  sensitized  mice  to  light.  Sufficient  doses  of  hematopor- 
phyrin may  sensitize  mice  so  that  they  become  narcotized  and  die  in  a 
few  minutes  after  exposure  to  intense  light,  a  true  "light  stroke." 

Pseudomelanosis. — When  loosely  bound  iron  is  present  in  the  tis- 
sues, and  in  the  same  tissues  sulphides  are  produced  through  bacterial 
action,  a  discoloration  with  sulphide  of  iron  will  result,  which  is 
'Called  pseudomelanosis,  because  the  pigment  resembles  true  melanin 
in  its  blackness.  This  is  most  frequently  observed  as  a  postmortem 
phenomenon  in  and  about  the  abdominal  cavity,  and  in  the  ordinary 
postmortem  discoloration  both  the  liberation  of  the  iron  from  its  firm 
organic  combination,  and  the  production  of  hydrogen  sulphide,  are  the 
■work  of  bacteria.  Pseudomelanosis  may  also  occur  intra  vitam,  par- 
ticularly in  the  margins  of  infected  areas,  and  it  may  also  be  observed 
in  the  intestines,  liver  and  spleen,  and  about  the  peritoneum,  in  bodies 
examined  immediately  after  death,  before  any  evident  postmortem 
decomposition  has  set  in.  This  seems  to  depend  upon  the  previous 
intra  vitam  formation  of  hemosiderin,  which  is  then  combined  by  sul- 
phur liberated  from  tissue  proteins  through  bacterial  action.''^  If 
hj'drogen  sulphide  acts  upon  hemoglobin  that  has  not  been  decom- 
posed, a  greenish  compound  of  sulphur-mctheniogloTjin  is  formed 
(Harnack^^),  which  is  the  cause  of  the  greenish  color  seen  in  the 
abdominal  walls  and  along  the  vessels  of  cadavers.  This  union  of 
hemoglobin  and  hydrogen  sulphide  occurs  only  when  oxygen  is  pres- 
ent (oxyhemoglobin).  The  sulphur-hemoglobin  compound  is  readily 
decomposed  by  weak  acids,  even  by  CO.,  with  the  formation  of 
methemoglohin,  which  in  turn  readily  becomes  decomposed  to  form 
hematin. 

During   life   sulphemoglohin  may   form,   the   sulphur  presumably 

coming  from  ijitestinal  putrefaction,  and  hence  called  "enterogenous 

cyanosis,"  which  term  also  covers  metliemoglohinemia  produced  by 

nitrites  formed  in  the  intestines.^"     The  latter  condition  is  also  pres- 

67a  Dent.  Arch.  klin.  Med.,  191.3   (112).  476. 
G8  Ernst,  Virchow's  Arch.,   189S    (152),  418.     Literature. 
69Zeit.  physiol.  Chem.,  1899   (20),  .558. 

70  West  and  CMarke.  Lancet.  Feb.  2,   1907:   Davis,  ihid.,  Oct.  26,   1912;   Gibson 
Quart.  Jour.  Med.,  1907   (1),  29;  Wallis,  Hid.,  Oct.,  1913. 
31 


482  I'ATIIOLOaiCAL    PldMEXTATION 

ent  in  poisoning  by  plienacetin,'"'^  aniline  and  acetanilid,  and  related 
pigments  appear  in  the  blood  in  poisoning  with  chlorates  and  uitro- 
benzol.  Pneumocoeei  and  ^Streptococcus  viriduns,  as  well  as  some 
other  bacteria,  may  produce  methemoglobin.^'"'  In  infections  with 
B.  empliysematosus,  Scluimm  found  this  pigment  free  in  the  blood, 
and  probably  it  could  be  found  in  other  conditions  if  sought. 

Hemofuscin  is  the  name  given  by  von  Recklinghausen  to  the 
brownish  pigment  found  in  involuntary  muscle-tibers,  particularly  in 
the  wall  of  the  intestine.  It  does  not  react  for  iron,  and  is  insoluble 
in  alcohol,  ether,  chloroform,  or  acids;  therefore  it  is  not  a  iipochrome. 
It  is  bleached  by  HoOo,  and  is  often  found  associated  with  hemosiderin 
which  is  not  bleached.  Von  Recklinghausen,  and  also  Goebel,^^  ascribe 
this  pigment  to  an  alteration  of  hemoglobin  which  enters  the  cells  in 
dissolved  form,  but  Rosenfeld,'-  who  has  submitted  the  material  to 
analysis  after  isolation,  found  3.70  per  cent,  of  sulphur,  from  which  he 
considers  that  it  is  related  to  the  melanins  or  melanoid  substances. 
The  substance  is  readily  dissolved  by  alkalies,  and  contains  no  iron. 
According  to  Taranoukhine,^^  the  pigment  in  the  myocardium  in 
hroioi  atrophy  of  the  heart  is  also  derived  from  proteins,  and  is 
neither  a  Iipochrome  nor  a  hemoglobin  derivative.  Other  observers, 
however,  consider  this  pigment  a  Iipochrome  or  a  lipofusein.  It  is 
probable  that  the  name  hemofuscin  has  been  given  to  several  different 
pigments,  which  resemble  one  another  only  in  that  they  do  not  contain 
iron.  Strater  "^'^  says  that  the  name  hemofuscin  cannot  be  used  for 
the  pigment  of  the  involuntary  muscles,  as  he  finds  evidence  that  it 
does  not  arise  from  hemoglobin  and  is  probably  a  waste  pigment :  but 
hemofuscin  is  found  in  e}>ithelial  and  connective  tissue  cells. 

Hemochromatosis.'* — This  name  was  given  by  von  Recklinghausen 
to  a  condition  in  which  the  organs  and  tissues  throughout  the  body  are 
abundantly  infiltrated  with  two  pigments :  one,  iron-containing,  iden- 
tical with  hemosiderin ;  the  other  seems  to  be  the  same  as  the  hemo- 
fuscin described  above.  It  is  to  be  distinguished  from  general  hemo- 
siderosis in  which  only  the  iron  pigment  is  deposited.'*''  In  hemo- 
chromatosis the  hemosiderin  is  found  chiefly  in  the  parenchyma  cells 
of  the  glandular  organs,  especiall.y  the  liver  and  pancreas,  which  or- 
gans usually  show  marked  interstitial  proliferation.     Hess  and  Zur- 

TOaRee  PTciibner.  Arch.  exp.  Path.,  1913    (72),  241. 

70b  CoU'.  Jour.  Exp.  ;Med..  1914  (20).  .303;  Blake  ihid..  1910  (24).  315; 
Sohumni,  Zoit.  phvsiol.  Ciicm.,  1913    (87),  171. 

Ti  Virchow's  Arch.,  1894    (130),  4S2. 

"Arch.  exp.  Path.  ii.  Pharm..  1900   (45),  40. 

"Rousskv  Arch.  Patol.,  1900    (10),  441. 

-3a  Vircliow's  Arch.,  1914    (218),   1. 

74  Literature  piven  by  Sprunt,  Arch.  Int.  Med.,  1911  (8).  75;  Potter  and 
Milne,  Aincr.  Jour.  Med!  Sci.,  1911  (143),  46;  Roth,  Deut.  Arcli.  kiln.  Mod..  1915 
(117),  224. 

7+a  In  lower  animals  occurs  a  form  of  liemochromatosis  afTectinir  especially  the 
bones,  and  sometimes  mistaken  for  ochronosis.  (See  Teutschlaender  X'irchow's 
Arch.,    1914     (217),    393.) 


iii:M<)('/ih'<)\iAT0Sis  483 

lu'lle  i'oniul  ;)S.7  ^in.  of  iron  in  llic  livci-  in  one  case  (the  normal 
amount  is  0.3  gm.),  and  Bemouille  "  found  18.3  gm.  or  2,95  per  cent, 
of  the  dry  weifrht  in  the  liver,  2.65  per  cent,  in  the  pancreas,  and  the 
same  in  tlie  spleen.  Anschiitz  found  14.69  per  cent,  in  the  lymph 
glands,  7.62  per  cent,  in  the  liver,  and  5  per  cent,  in  the  pancreas  of 
a  case,  ^hi'ir  and  Dunn  "''  obtained  the  following  percenttige  figures : 
Liver,  6.43 ;  pancreas,  2.49 ;  spleen,  0.825 ;  retroperitoneal  glands, 
11.64;  kidneys,  0.406:  adrenals,  0.121;  heart,  0.714;  skin,  0.188; 
small  intestine,  0.14.  The  hemofuscin  is  found  in  the  smooth  muscle- 
fibers  of  the  gastro-intestinal  tract,  blood-vessels,  and  genito-urinary 
tract.  Under  the  heading  of  local  hemochromatosis,  von  Reckling- 
hausen grouped  such  conditions  as  brown  atrophy  of  the  heart,  and 
pig-mentation  of  the  intestinal  wall,  which  probably  are  quite  dis- 
tinct from  the  generalized  hemochromatosis,  since  the  local  form  oc- 
curs as  a  physiological  process  in  old  age. 

In  a  considerable  proportion  (50  of  63  collected  by  Sprunt)  of  the 
cases  of  generalized  hemochromatosis  there  occurs  diabetes,  called  by 
Hanot,  "bronzed  diabetes,"  because  of  the  coloration  of  the  skin. 
It  has  been  suggested  that  the  pigmentation  is  due  to  decomposition 
of  the  blood-corpuscles  in  the  diabetic  blood,  but  the  pigmentation 
and  sclerotic  changes  precede  the  diabetes,  which  is  secondary  to  the 
atrophic  and  sclerotic  changes  in  the  pancreas.  There  can  be  little 
question  that  both  the  pigment  formation  and  the  tissue  changes  de- 
pend upon  some  intoxication,  the  origin  and  nature  of  the  toxic  agent 
being  entirely  unknown.  In  many  cases  it  has  seemed  probable  that 
alcohol  might  have  been  the  inciting  cause.  There  is  no  evidence  of 
any  abnormal  blood  destruction  which  might  account  for  the  pigmen- 
tation, and  Parker  suggests  that  the  difficulty  lies  in  the  inability  of 
the  tissues  to  get  rid  of  the  iron  set  free  in  normal  catabolism.'^^'' 
Roessle  believes  that  the  primarv^  change  is  in  the  capillaries,  whereby 
hemorrhagic  extravasations  take  place,  and  phagocytosis  of  red  cor- 
puscles by  gland  cells  results  in  pigmentation. 

Opie's  conclusions  concerning  this  subject  are  as  follows:  (1) 
There  is  a  distinct  morbid  entity,  hemochromatosis,  characterized  by 
widespread  deposition  of  an  iron-containing  pigment  in  certain  cells, 
and  an  associated  formation  of  iron-free  pigments  in  a  variety  of 
localities  in  which  pigment  is  found  in  moderate  amount  under  physi- 
ological conditions.  (2)  With  the  pigment  accumulation  there  occur 
degeneration  and  death  of  the  containing  cells  and  consequent  inter- 
stitial inflammation,  notably  of  the  liver  and  pancreas,  which  become 
the  seat  of  inflammatory  changes  accompanied  by  hypertrophy  of  the 
organ.  (3)  When  chronic  interstitial  pancreatitis  has  reached  a  cer- 
tain grade  of  intensity,  diabetes  ensues,  and  is  the  terminal  event  in 

T5Corr.-Bl.  Scliweiz.  Aertze,   1911    (40).  010. 
Tsa.Jour.  Path,  and  Ract.,  1914   (19),  226. 
-EbSee  Quart.  Jour.  Med.,  1914    (7),  129. 


484  PATHOLOGICAL    PIGMEXTATION 

the  disease.  Spmnt  suggests  that  the  diabetes  may  be  referable  to 
diminished  oxidative  power  because  of  disturbances  in  the  iron-con- 
taining constituents  of  the  tissues,  assuming  these  iron  compounds  to 
be  catalytic  agents  in  oxidizing  processes. 

ICTERUS  ■« 

Pigmentation  of  the  tissues  of  the  hody  in  jaundice  depends  upon 
the  presence  in  them  of  bile-pigments,  which  usually  have  been  formed 
in  the  liver  and  reabsorbed  either  into  the  lymph  or  blood  (or  both). 
However,  a  pigment  that  seems  to  be  chemically  identical  with  bili- 
rubin {hematoidin)  may  be  formed  from  hemoglobin  liberated  on  the 
breaking  up  of  red  corpuscles,  and  possibly  this  may  be  produced  in 
sufficient  amounts  outside  of  the  liver  to  give  rise  to  general  icterus. 
Certainly  the  local  greenish-yellow  pigmentation  occurring  in  the 
vicinity  of  extravasations  of  blood,  due  to  hematoidin  formation,  may 
be  looked  upon  as  a  "local  jaundice. "^^  and  in  icterus  hematoidin 
crystals  may  be  found  in  the  tissues." 

Bile-pigments. — Biliruhin  is  of  a  reddish-yellow  color,  and  it  is  the  chief  pig- 
ment of  human  hile.  Its  formula  is  Cg^H^sNiOu  or  C.,3H3oN40,;,  and  its  relation  to 
hematin,  from  wliich  it  is  formed,  is  sliown  by  the  following  formula,  which  ex- 
presses the  manner  in  which  blood  pigment  may  Ijc  converted  into  bilirubin  by 
the  liver  under  normal  conditions,  and  into  hematoidin  (its  isomer)  in  the  tissues 
and  fluids  of  the  body  in  pathological  conditions: 

Ca^H^^N.O.Fe  +  2HoO  =  C^JI.s'^fi,  f  FeO. 
(hematin)  (hematoidin  or 

bilirubin) 

Bilirubin  is  not  soluble  in  water,  but  dissolves  in  the  alkaline  body  fluids  as  a 
soluble  compound,  "bilirubin  alkali."  It  is  very  slightly  soluble  in  ether,  ben- 
zene, carbon  disulphide,  amyl-alcohol,  fatty  oils,  and  glycerol,  Init  is  more  soluble 
in  alcohol  and  in  chloroform. 

Biliverdin,  C;,4H38N40s,  as  its  formula  indicates,  is  an  oxidation  product  of 
bilirubin.  Bilirubin  in  alkaline  solutions  will  oxidize  into  biliverdin  merely  on 
exposure  to  tlie  air,  and  the  change  from  yellow  to  green  of  icteric  specimens  when 
placed  in  oxidizing  solutions  (e.  g.,  dicliromate  hardening  fluids)  is  due  to  the 
formation  of  the  green  biliverdin.  Biliverdin  is  the  chief  pigment  of  tlie  bile  of 
carnivora,  but  it  is  also  present  in  varying  amounts  in  human  bile. 

The  various  other  biliary  pigments,  namely,  hilifuscin,  hiliprdsin.  rliolrprnniii.'^ 
hilihininn,  and  hilicyanin,  are  probably  not  normal  C(mstituents  of  bile,  but  are 
oxidation  products  of  bilirubin,  and  are  found  chiefly-  in  gall  stones  ( (/.  c. ) .  A 
pigment  similar  to  urobilin  may  be  present  in  normal  bilo.  The  total  amount  of 
pigment's  present  in  liile  is  probably  not  far  from  one  gram  per  liter;  ratlier  under 
than  above  this  amount. 

Etiology  of  Icterus. — Although  hematoidin,  wliich  is  isomeric  if 
not  identical  with  bilirubin,  may  be  formed  outside  of  the  liver  when 
red    corijuscles    are    broken    u])    in    hemoiThagic    extravasations,    and 

.  76  Literature  by  Stadelmann,  "Der  Icterus,"  Sluttgart,  1S91;  Minkowski, 
Ergebnisse  der  Pathol.,   1895    (2),  079. 

77  See  Ouillain  and  Troisier,  Semaine  MM.,  1909  (29).  l;?:5:  Widul  and  .Jolt rain, 
Arch.  mr-d.  expC'r.,  1909   (21).  (541. 

78l)iinzelt,  Cent.   f.  Patli.,    1909    (20).   900. 

70  See  Kiister,  Zeit.  physiol.  Chem.,   1906    (47),  294. 


ICTERUS  485 

possibly  also  when  they  are  broken  up  within  the  vessels  by  hemolytic 
agents,  yet  it  has  generally  been  considered  that  a  true  general  icterus 
does  not  occur  without  the  liver  being  implicated.  This  view  rested 
on  evidence  of  various  sorts.  First,  the  classical  experiments  of 
^Minkowski  and  Nannyn,"^'  which  demonstrated  that  in  geese  the  pro- 
duction of  hemolj-sis  by  means  of  arseniuretted  hydrogen  leads  to 
icterus,  but  if  the  livers  of  the  geese  have  been  previously  removed, 
no  icterus  follows  the  poisoning.  Second,  the  repeated  demonstration 
that  in  icterus  produced  by  septic  conditions,  poisoning,  etc.,  which 
was  formerly  looked  upon  as  a  ''hematogenous"  icterus,  the  urine 
contains  bile  salts  as  Avell  as  pigment,  indicating  an  absorption  of  bile 
from  the  liver.  Third,  the  finding  of  histological  evidence  that  in 
so-called  hematogenous  icterus  there  occur  occlusions  or  lesions  of 
some  sort  in  the  bile  capillaries,  which  can  account  for  the  reabsorp- 
tion  of  the  bile  into  the  general  circulation.®^  Therefore,  it  was  be- 
lieved that  the  pigments  that  produce  the  general  discoloration  of 
icterus  are,  at  least  for  the  most  part,  manufactured  by  the  liver, 
whatever  the  cause  of  the  reabsorption  of  the  bile  from  the  liver  into 
the  blood  may  be.  That  hemolytic  agents  cause  icterus  was  explained 
by  the  fact  that  on  account  of  the  large  amounts  of  free  hemoglobin 
brought  to  the  liver,  excessive  amounts  of  bile-pigments  are  formed, 
which  render  the  bile  so  viscid  that  it  blocks  up  the  fine  bile  capil- 
laries; on  account  of  the  low  pressure  at  which  bile  is  secreted,  a 
slight  obstruction  of  this  kind  is  sufficient  to  stop  entirely  the  outflow 
of  bile,  which  then  enters  the  capillaries  of  the  liver  and  also,  to  a  less 
extent,  the  lymphatics.^^  It  is  also  possible  that  the  hemolytic  poisons 
injure  the  liver-cells  so  much  that  the  minute  intra-  and  intercellular 
bile  capillaries  become  disorganized,  and  permit  of  escape  of  bile  into 
the  lymph-spaces  and  its  absorption  into  the  blood-vessels.®^  Swelling 
of  the  degenerated  liver-cells  may  also  be  an  important  factor  in  the 
occlusion  of  the  bile  capillaries;  swelling  of  the  lining  cells  of  the  bile 
capillaries  may  also  coexist,  and  fibrin  may  occlude  them  in  toxic  or 
infectious  icterus. 

However,  Whipple  and  Hooper,®*  have  obtained  experimental  evi- 
dence that  after  intravenous  injection  of  hemoglobin  into  dogs  with 
the  liver  excluded  from  the  circulation,  bile  pigments  appear  in  the 
urine  and  icterus  is  manifested  in  the  fat  tissues,  from  which  observa- 
tions it  is  concluded  that  the  liver  may  not  be  the  only  place  in  which 

80  Arch.  f.  exp.  Pathol,  u.  Pharm.,  1886    (21),  1. 

81  See  Eppin^er,  Ziegler's  Beitr..  1903  (33),  123;  Gerhardt,  Miindi.  med.  Wooh., 
1905  (52),  889.  Lang  (Zeit.  exp.  Path.  u.  Ther.,  July,  1906  (3),  473)  has 
demonstrated  the  presence  of  fibrinogen  in  the  bile  in  phosphorus-poisoning,  which 
perhaps  accounts  for  the  "l)ile  thrombi"  observed  by  Eppinger  in  toxic  icterus. 

82  See  :Mendel  and  Underbill,  Amer.  Jour.  Phvsiol.,  1905  (14),  252;  Whipple 
and  King,  Jour.  Exp.  ]\red.,   1911    (13),   115. 

s'?  Sterling,  Arch.  exp.  Path.,  1911  (64),  468;  Fiessinger,  Jour.  Phvsiol,  et 
Pathol.,  1910    (12),  958. 

84  Jour.  Exper.  Med.,  1913    (17),  593  and  612. 


486  PATHOLOGICAL    PIGMEXTATION 

bile  pigment  can  be  formed  from  hemoglobin.  Several  authors  have 
found  bilirubin  produced  in  hemorrhagic  elfusions  located  where  the 
liver  could  have  had  no  influence.®^''  We  also  recognize  types  of  hemo- 
lytic icterus  in  which  the  liver  does  not  seem  to  be  concerned,  and 
with  bile  pigments  present  in  the  blood  and  urine  unaccompanied  by 
bile  salts  (dissociated  icterus),  so  that  the  old  dictum  of  the  essential" 
implication  of  the  liver  in  icterus  seems  to  be  incorrect.^*''  Joanno- 
vics  "  gives,  as  a  result  of  a  comparative  study  of  icterus  from  bile 
obstruction  and  icterus  from  hemolysis,  the  following  chief  differ- 
ences: Icterus  due  to  hemolysis  appears  sooner  than  icterus  from 
bile-duct  occlusion,  and  reaches  a  much  higher  degree ;  the  obstruction 
in  hemolytic  icterus,  Avhen  present,  is  intra-acinous ;  in  stasis  it  is 
cliiefly  inter-acinous ;  in  hemolytic  icterus  there  is  a  large  splenic 
tumor  due  to  accumulation  of  degenerated  red  cells  in  the  spleen, 
where  they  become  disintegrated  preliminary  to  the  formation  of  bile- 
pigment.  If  the  spleen  is  removed,  hemolytic  agents  may  not  cause 
icterus,  because  the  corpuscles  are  not  then  prepared  for  pigment 
formation. ^^-'^ 

Toxicity  of  Bile. — In  any  event,  we  must  appreciate  that  in  icterus 
not  only  are  abnormally  large  quantities  of  bile-pigment  present  in 
the  blood,  but  also  the  other  less  conspicuous  constituents  of  the  bile. 
Whole  bile  of  rabbits  is  fatal  to  rabbits  in  doses  of  0.25  to  0.5  cc.  per 
kilo,  by  intraperitoneal  injection,  and  about  half  as  much  intraven- 
ously (Bunting  and  Brown  ^'^).  Death  is  the  result  of  changes  in  the 
myocardium,  where  necrosis  is  produced ;  and  severe  degenerative 
changes  are  also  found  in  the  kidneys  and  liver;  when  the  bile  is  in- 
jected into  the  peritoneum,  pancreatitis  and  fat  necrosis  result.  The 
relative  toxicity  of  the  bile-pigments  and  the  bile  salts  is  not  as  yet  uni- 
formly agreed  upon. 

Bile-pigments. — Bouchard  *^  and  others  have  claimed  that  the  bile- 
pigments  are  far  more  toxic  than  the  bile  salts,  which  is  contradicted 
by  Rywosch  and  others.  A  series  of  analyses  by  Gilbert  '^'"^  and  others 
gave  the  following  results :  Normal  blood-serum  contains  0.027-0.08 
gram  bilirubin  per  liter;  in  obstructive  ictenis  they  found  0.7  to  1.0 
gram  of  bilirubin  per  liter,  in  biliary  cirrhosis  0.33  gram  per  liter,  in 
icterus  neonatorum  0.2  to  0.5  gram ;  in  pneumonia  0.068  gram  was 
found.     King  and  Stewart  -^  state  that  the  amount  of  pigment  in  a 

s<4aTIoop(M-  and  ^^■llippl(>,  Jour.  Exp.  Mod.,  inifi   (2:1),  137. 

S4b  Att('iiij)ts  to  produce  hile  pifjments  from  liemojiloliin  by  bacterial  action 
have  been  un.successful.      (Quadri,   Fol.  Clin.  C'liim..  1914,  Xo.  10). 

ssZeit.  f.  Hellk.,  Path.  Al)t..  l')()4   (25),  25. 

S5a 'I'lic  etiology  of  icterus  neonatorum  (when  not  obstructive)  has  not  been 
ascertained,  but  a  nalurai  tendency  towards  icterus  is  said  to  exist  in  tiie  new- 
l)orn,  llieir  l)b>od  containin<r  much  more  bile  ]u<jment  tlien  than  later,  (llirsch. 
Zeit.  Kinderheilk.,  ]it]3   (0),  10(1;  Ylppo,  Miinch.  med.  Woch.,  101,3    (39),  2161.) 

^«.7our.   Kxper.  Med.,   mil    (14),  445. 

f+T  Literature  and  discussion  by  StadelTiiaiui.  Zeit.  f.  V,\o\.,   1806    (34),  57. 

«7aC"ompt.  Rend.  Soe.  Biol.,   1005  and   1006. 

xs.Tour.  Exper.  ISled.,   1000    (11),  673. 


ICTERUS  487 

Icllial  (lose  of  bile  will  cause  death,  but  the  bile  salts  present  in  the 
same  (juantity  of  bile  will  not  cause  recognizable  effects;  unconibined 
pigment  is  more  toxic  than  its  calcium  or  magnesium  salts.  Bile  from 
which  the  pigment  is  removed  has  very  little  toxicity.  They  suggest 
that  calcium  is  increased  in  the  blood  in  icterus  as  a  protection  against 
the  toxic  effects  of  the  pigments.  The  combining  of  the  calcium  with 
bile  pigment,  however,  renders  it  unavailable  for  fibrin  formation,  and 
this  seems  to  be  an  important  factor  in  the  hemophilic  tendency  of 
icterus.**^''  The  decrease  in  available  calcium  may  also  be  responsible 
for  the  bradycardia  and  some  of  the  mental  and  nervous  symptoms. 

Bile  salts  are  undoubtedly  toxic,  generally  producing  depression 
of  the  central  nervous  system,  with  resulting  coma  and  paralysis; 
they  are  also  decidedly  toxic  to  cells  of  all  sorts,  causing  hemolysis  and 
marked  destiTiction  of  tissue-cells.  Small  quantities  of  bile  salts 
stimulate  the  central  end  of  the  vagus,  and  large  amounts  influence 
the  heart  itself;  hence  in  icterus  we  observe  a  slowing,  and  often  an 
irregularity,  of  the  pulse,  and  the  blood  pressure  is  lowered.  Al- 
though there  has  been  much  dispute  as  to  whether  the  chief  effects  of 
icterus  upon  the  heart  depend  upon  action  of  the  bile  salts  upon  the 
vagus,  or  upon  the  intracardiac  ganglia,  or  upon  the  muscle  itself,^" 
yet  Weintraud  demonstrated  that  in  some  cases  of  icterus  administra- 
tion of  atropin.  which  paralyzes  the  vagus,  stops  the  bradycardia, 
indicating  the  importance  of  the  effects  of  the  bile  salts  upon  the 
vagus  in  causing  this  feature  of  cholemia.  According  to  ^leltzer  and 
Salant,"°  bile  also  contains  a  tetanic  element,  which  disappears  from 
stagnating  bile ;  the  bile  salts  contain  this  tetanizing  agent  in  less 
amount  than  does  the  whole  bile.  But  King  ^^  and  others  ascribe  most 
of  the  effects  of  bile  on  the  heart  to  the  bile  pigments,  perhaps  through 
abstraction  of  the  calcium. 

Since  the  bile  salts  cause  hemolj^sis,  and  since  in  even  "hematogen- 
ous" jaundice  they  may  enter  the  blood,  it  can  readily  be  seen  that  in 
this  way  an  increased  formation  of  bile-pigment  may  be  incited  which 
leads  to  further  obstiiiction  to  the  outflow  of  bile  from  the  liver,  and 
a  "vicious  circle"  maj-  thus  be  established.  The  necroses  observed  in 
the  liver  in  icterus,  "icteric  necrosis,"  are  generally  ascribed  to  the 
cytotoxic  effects  of  the  bile  salts,  although  it  is  difificult  always  to 
exclude  infection  extending  along  the  bile-ducts  to  the  liver  tissue. 
Possibly  the  power  of  bile  salts  to  dissolve  lipoids  may  be  responsible 
for  the  cytotoxic  effects "-  as  well  as  for  the  hemolysis.  The  itching 
and  irritation  of  the  skin  in  icterus  may  be  due  to  the  effect  of  the 
bile-salts  deposited  in  it,  for  pruritis  is  said  to  be  absent  in  the  pig- 

88a  See  Lee  and  Vincent,  Areli.  Int.  :Med.,  10]5    (16),  59. 

89  See  Minkowski.  Erpeb.  der  Patiiol.,  lSf>5    (2),  709. 

90  Jour.  Exp.  Med.,  1906  (8),  128;  review  and  literature  concerning  toxicity  of 
bile. 

91  See  King,  Bigelow  and  Pearce,  Jour.  Exper.  Med.,  1912   (14),  159. 
92Neufeld  and  Hiindel,  Arb.  kaiserl.  Ges.-Amte,  1908   (28),  572. 


488  PATHOLOGICAL    PIGMENTATION 

mentary  jaundice  of  congenital  hemolytic  icterus.  There  is  also  an 
increase  in  the  cholesterol  in  the  blood,  which  may  be  related  to  the 
"xanthomas"  that  form  in  chronic  icterus."^ 

A  remarkable  tendency  to  spontaneous  hemorrhages,  frequently  ob- 
served in  icterus,  probably  depends  upon  injury  to  the  capillary 
endothelium  by  the  bile  salts,"^  while  the  protracted,  often  uncontrol- 
lable, hemorrhage  that  may  occur  from  operation  wounds  in  icteric  pa- 
tients, is  related  to  the  slowed  coagulation  of  the  blood  observed  in 
icterus.  The  cytotoxic  effect  of  the  bile  salts  is  also  shown  by  the 
albuminuria  of  icteric  persons,  which  frequently  results  from  the 
renal  lesions  the  bile  produces.  Although  bile  itself  is  toxic  to  many 
bacteria,  especially  the  pneumococcus,^^  yet  in  icterus  the  bactericidal 
power  of  the  blood  is  lowered,  and  infections  are  prone  to  develop  and 
to  be  severe;  moreover,  the  growth  of  several  species  of  bacteria  is 
favored  by  bile."*^ 

Croftan  ^^  summarizes  the  physiological  effects  of  bile  acids  as  fol- 
lows: (1)  A  powerful  cytolytic  action,  affecting  both  blood-cor- 
puscles and  tissue-cells.  (2)  A  distinct  cholagogue  action.  (3)  In 
small  doses  (1-500)  they  aid  coagulation.  (4)  In  large  doses  (1-250 
and  over)  they  retard  coagulation.  (5)  Slow  the  heart  action.*'^'^ 
(6)  In  small  doses  they  act  as  vasodilators;  in  large  doses,  as  vasocon- 
strictors. (7)  Reduce  motor  and  sensory  irrita-bility.  (8)  Act  on 
the  higher  cerebral  centers,  causing  coma,  stupor,  and  death.  Sel- 
lards  °*  found  that  injection  of  bile  salts  into  guinea  pigs  causes  ulcer- 
ation and  hemorrhage  in  the  stomach. 

It  is  difficult  to  decide  how  much  of  the  profound  intoxication  that 
is  sometimes  present  in  icterus  (''cholemia"  and  "icterus  gravis") 
to  ascribe  to  the  reabsorbed  bile,  for  frequently  there  is  an  accompany- 
ing infection,  and  even  if  there  is  no  infection  the  impairment  of  liver 
function  by  the  obstruction  of  bile  outflow  must  also  be  reckoned 
with.  The  liver  is  not  only  the  great  destroyer  of  toxic  substances 
absorbed  from  the  alimentary  canal,  but  it  is  also  an  important  seat 
of  nitrogenous  metabolism,  interference  with  which  may  lead  to  ac- 
cumulation of  many  toxic  nitrogenous  substances  in  the  blood. ^  The 
long  duration  of  severe  icterus  in  some  cases  of  occlusion  of  the  bile- 
ducts,  with  relatively  slight  evidences  of  intoxication,  would  seem  to 
indicate,  however,  that  on  the  whole  the  bile  is  not  so  nuich  respon- 
ds ChaufTard,  Prosso  Mr>d.,  1913  (21),  SI;  Chvostok.  Zcit.  klin.  Med.,  1!)11  (73), 
470;  Pinkiis  and  Pick,  Dout.  med.  Woch.,  1008    (34),  1427. 

n-tSpo  Morawitz,  Areh.  cxp.  Path..    1907    (56),   115. 

"•'>  Roe  Noiifpld  and  TTaendpl,  Inr.  cit. 

n«  See  Mcyorstoin,  fVnt.  f.  Pnkt..  1907    ^44),  434. 

f'7New  York  Mod.  .Tour.,  inn;  (;.3).  SIO;  see  also  Faust,  "Die  tiorisohe  Cifto," 
Braunschwoifr,  1906,  p.  29. 

oTaSoo  Porti,  Oaz.  depli  Ospod..  1910    (37),  1233. 

!>«Aroli.  Tnt.  Mod.,  1909    (4),  .'>02. 

1  See  Bickel,  Exper.  Untersuch.  liber  der  Pathol,  der  Cholaemie,  Wiesbaden, 
1900. 


CONGENITAL  HEMOLYTIC  ICTERUS  489 

sible  for  the  iiitoxieation  observed  in  icterus  as  are  the  associated 
conditions.  On  the  other  hand,  in  not  a  few  instances  it  has  been 
observed  that  escape  of  large  quantities  of  bile  into  the  peritoneal 
cavity  may  be  followed  by  symptoms  similar  to  those  of  icterus  gravis; 
in  these  cases  only  the  bile  can  be  held  responsible  for  the  intoxica- 
tion.- 

Dissociated  Jaundice -i^  is  the  oxistcneo  of  cillicr  l)ilc  salts  or  l)ilo  pi<j;ment  sep- 
arately in  the  blood.  This  may  be  produced  either  by  the  bile  salts  beinjr  ex- 
creted by  the  kidney,  leaving  only  the  le^s  dillusible  piprinent  in  the  blood,  or  by 
separate  escape  of  bile  salts  from  the  liver  into  the  blood.  Also  in  true  liem- 
olytic  icterus  we  may  have  bile  pigments  present  in  the  blood  without  bile  salts. 

CONGENITAL  HEMOLYTIC  ICTERUS  2a 

This  term  describes  a  cimdition  charat'teri/ed  by  a  chronic,  non-obstructive 
jaundice,  without  evident  intoxication.  A  similar  condition  is  also  obser\ed  de- 
veloping in  adults,  without  familial  tendencies.  The  congenital  form  usually 
shows  familial  character,  but  isolated  congenital  cases  do  occur.  It  is  the  re- 
sult of  active  hemolysis,  apparently  taking  place  chiefly  in  the  spleen,  and  lead- 
ing to  an  icterus  without  evident  participation  of  the  liver.  Tlie  cause  of  the 
hemolysis  is  entirely  unknown,  althougli  there  is  a  marked  fragility  of  the 
erythrocytes  evidenced  by  reduction  of  their  resistance  to  hypotonic  solutions,  and 
it  results  in  a  moderate  anemia,  with  excretion  of  much  urobilin  in  l)oth  stools 
and  urine;  the  blood  contains  bilirubin  wliich  is  not  excreted  in  the  urine.  The 
jaundice  is  usually  unaccompanied  by  evidences  of  cholemia.  icteric  pruritis  or 
hemophilia.  The  spleen  is  greatly  enlarged  and  improvement  has  generallj'  fol- 
lowed splenectomy  but  the  exact  relation  'of  the  spleen  to  tlic  disease  is  not 
known. 2b  The  frequent  occurrence  of  gall  stones  in  this  condition  may  be  the  re- 
sult of  hypercholesterolemia  from  hemolysis. 

The  metabolism  of  a  case  2c  showed  loss  of  nitrogen,  calcium,  maenesium  and 
iron,  and  a  much  increased  uric  acid  excretion.  These  conditions  may  improve 
after  operation. 2d 

The  Pigmentation  in  Icterus. — Living  tissues  have  but  a  slight 
tendency  to  take  up  bile-pigments,  much  of  the  tissue-staining  ob- 
served at  autopsy  being  due  to  postmortem  imbibition  from  the  blood 
and  lymph.  Quincke  ^  found  that  after  .subcutaneous  injection  of 
bilirubin  only  the  connective  tissue,  both  cells  and  intercellular  fibrils, 
becomes  diffusely  colored ;  later,  it  fades  out  of  the  cells,  leaving  only 
the  fibrils  stained.  ]\Iuscle-cells,  fat-cells,  and  vessel-walls  take  up  the 
pigment  only  after  their  death.  If  the  jaundice  continues  for  a  long 
time,  the  subcutaneous  deposits  of  bilirubin  may  undergo  a  slow  oxida- 
tion, the  color  changing  to  an  olive  or  to  a  dirty  grayish  green.  The 
pigment  in  the  connective  tis,sues  is  at  first  in  solution,  but  may  be  de- 
posited in  a  granular  form  after  a  considerable  amount  has  accumu- 
lated. Bile  pigments  and  bile  salts  may  both  be  present  in  consider- 
able amounts  in  the  blood  and  not  pass  through  the  kidneys,  and  also 

2  See  Ehrhardt.  Arch.  klin.  Chir.,   1901    (64),  314. 
2e  Hoover  and  Blankenhorn.  Arch.  Int.  Med..  1010    (18),  289. 
2a  See  Richards  and  Johnson,  Jour.  Amer.  Med.  Assoc.,  191.3    (61),  1586. 
2b  See   series  of  articles  by  Pearce  ct  al.   in   Jour.   Exp.  ^led.,  on   Relation  of 
Spleen  to  Blood  Destruction. 

2c  IMcKelvy  and  Rosenbloom,  Arch.  Int.  Med..  IOI.t    (In),  227. 

2d  Goldschmidt,  Pepper  and  Pearce,  Arch.  Int.  Med.,  191.5    (16),  437. 

3Virchow's  Arch.,  1884    (95),  125. 


490  PATHOLOdlCAL    I'IGMEXTATION 

they  may  fail  to  pass  into  the  tissues;  hence  we  may  have  cholemia 
without  icterus  or  chohiria,  because  of  the  tirmness  with  wliich  tlie  pig- 
ments are  bound  in  the  plasma  (Hoover  -^). 

The  question  whether  in  icterus  the  skin  may  be  colored  by  other 
pigments  than  bilirubin,  especially  by  its  reduction  product,  urobilin, 
seems  to  have  been  decided  negatively.  Hile-pignient  is  probably  not 
absorbed  as  such  from  the  intestine  in  sufficient  quantity  to  cause 
icterus.  Such  bile-pigment  as  enters  tlie  blood  from  the  liver  is  ex- 
creted through  the  kidneys  chiefly,  but  also  in  the  sweat.  Ordinarily, 
other  secretions  (milk,  tears,  saliva,  sputum)  are  not  colored  in  jaun- 
dice, but  if  the  secretions  are  mixed  with  inflammatory  exudations, 
they  may  then  be  colored  (e.  g.,  pneumonic  sputum).  When  the  bile- 
pigment  is  resorbed  from  the  skin,  it  may  be  in  part  transformed  into 
urobilin,  which  appears  in  the  urine  in  increased  amounts  during  the 
period  of  recovery  from  jaundice.  Part  of  the  bile-pigment  is  prob- 
ably eliminated  by  the  liver  after  the  cause  of  obstruction  has  been 
removed  from  the  bile-passages. 

Urobilin  ^-'^  is  probably  formed  chiefly,  if  not  solely,  from  bile  pig- 
ments by  the  action  of  reducing  bacteria  in  the  intestine.  It  is  ex- 
creted in  the  urine  only  as  its  chromogen,  urobilinogen,  but  in  the 
feces  both  urobilin  and  urobilinogen  may  be  found ;  when  exposed  to 
air  the  chromogen  oxidizes  quickly  to  urobilin.  Addis  ^^  states  that 
bilirubin  is  reduced  to  urobilinogen,  in  the  bowel  and  is  then  largely 
absorbed,  to  be  at  once  oxidized  and  polymerized  into  urobilin,  two 
molecules  of  urobilinogen  uniting  under  the  influence  of  oxygen  to 
form  one  of  urobilin.  In  the  liver  the  urobilin  is  largely  worked  over 
to  form  new  hemoglobin,  and  hence  the  functional  capacity  of  the 
liver  is  indicated  by  the  completeness  with  which  it  utilizes  the  uro- 
bilin, except  in  cases  of  excessive  formation  of  urobilinogen  as  a  re- 
sult of  hemolysis.  The  amount  of  urobilinogen  in  the  urine  will  be 
found  increased,  therefore,  in  hemolytic  icterus,  and  decreased  in  ob- 
structive icterus.  However,  with  advanced  renal  disease  the  kidneys 
may  become  unable  to  excrete  the  urobilinogen  brought  to  them  in  the 
blood.  Exceptionally,  urobilinogen  may  be  fonned  from  blood  dis- 
integrated in  bloody  effusions  without  evident  participation  of  the 
liver,  e.  g.,  urobilinogenuria  in  hemorrhagic  ascites,  with  hemolytic 
poisons,  etc.  AVith  a  normal  liver  urobilinogeiuiria  is  found  only 
when  there  is  excessive  hemolysis,  otherwise  urobilinogenuria  occurs 
only  with  an  injury  to  the  liver  parenchynui  (Ilildebrant).  Occlu- 
sion of  the  bile  ducts  stops  an  existing  urobilinogenuria  by  prevent- 
ing the  formation  of  urobilinogen  in  the  intestine.  Normally  there  is 
a  very  small  amount  of  urobilinogen  and  related  substances  in  the 
urine,  which  disappears  when  there  is  no  bile  in  the  intestine.     From- 

"ii  T>il)lio<rrapliv  and  review  bv  INfever-Botz,  Erffpli.  inn.  ^lod.,   litl."]    (12).  734; 
Willnir  and  Addis,  Arcli.  Int.  Med.,  "l 014    (13),  235. 
ai^Arcli.   Fnt.  Med..   IOI.t    (15),  412. 


DI<!i:sTl\  E  DISTlU{B.\.yCE8  JX  OBSTRiCTIVE  ICTERUS  491 

holdt  ^'^  considers  that  increased  bacterial  reduction  in  the  intestines 
may  by  itself  account  for  urobilinogeniiria.  The  amount  of  urobilin 
and  urobilinogen  excreted  in  the  feces,  seems  to  vary  directly  with  the 
amount  of  hemolj'sis.^'* 

Digestive  Disturbances  in  Obstructive  Icterus.^ — In  tase  tlic  iotcnis  (Icpcnds 
upon  the  oochision  of  the  main  hilo-passajzos  by  stones,  tumors,  etc..  the  situation 
is  romplii'atod  l)y  tiu'  clloi'ts  of  the  ahsi-nci'  of  this  natural  secretion  in  the  in- 
testinal canal.  Carbohydrate  and  protein  dijjestion  seem  to  be  but  little  allected, 
especially  the  former,  but  the  proportion  of  the  ingested  fat  that  apj)ears  in  the 
feces  increases  from  the  normal  7-11  per  cent,  to  00-SO  per  cent.  The  products 
of  bacterial  decomjjositioii  of  the  undigested  fat  may  lead  to  injury  of  the  in- 
testinal wall  and  disturbance  of  its  function.  Failure  of  absorption  of  fat  also 
favors  intestinal  putrefaction  by  enveloj)ing  the  protein  substances  so  that  they 
are  not  readily  digested  and  absorbed.  The  relation  of  bile  to  intestinal  putre- 
faction is  still  not  exactly  determined.  Freiiuently,  but  by  no  means  always, 
ther<^  is  an  increased  intestinal  jnitrefaction  which  may  result  in  diarrhea  and 
the  appearance  of  excessive  quantities  of  indican  and  phenol  in  the  urine.  The 
idea  once  held  that  the  bile  salts  acted  as  intestinal  antiseptics  has  not  been 
establislied  by  experimental  investigations:  however,  it  is  possible  that  through 
their  fimction  as  natural  cathartics,  liy  stimulation  of  peristalsis,  they  prevent 
stagnation  and  putrefaction  of  proteins. 

3cZeit.  exp.  Path.,  1911   (9),  268. 
3d  Robertson,  Arch.  Int.  Med.,  191.5    (15),  1072. 

4  Concerning  metabolism  in  icterus  see  Vannini,  Zeit.  klin.  Med.,  1912  (75), 
136. 


CHAPTER   XVII 
THE  CHEMISTRY  OF  TUMORS  ' 

Chemical  investigations  of  tumors  have  been  relatively  few  in  num- 
ber, but,  so  far  as  they  have  yet  been  made,  there  has  been  detected 
little  that  indicates  any  important  deviation  of  the  chemical  processes 
of  tumors  from  those  of  normal  cells  of  similar  origin.  Likewise,  the 
chemical  composition  of  tumor  tissue  resembles  closely,  on  the  whole, 
the  composition  of  related  normal  tissues.  It  is  hardly  to  be  im- 
agined that  the  course  of  chemical  changes  is  greatly  different  in 
tumor  cells  from  that  in  normal  cells,  in  view  of  the  abundant  evi- 
dence that  the  metabolic  products  of  tumor  cells  are  identical  with 
those  of  the  cells  from  which  they  arose.  Thus,  metastatic  growths 
of  thyroid  tissue  will  produce  thyroiodin  in  any  part  of  the  body,  liver 
carcinoma  metastases  produce  bile,  tumors  from  the  choroid  or  from 
pigmented  moles  produce  melanin,  etc."  The  capacity  of  tumor  cells 
to  produce  complicated  products  of  metabolic  action  specific  for  the 
parent  cells  from  which  they  arose,  as  illustrated  above,  indicates 
bej^ond  question  that  the  coui*se  of  their  chemical  activities  is  very 
much  like  that  of  normal  cells.  So,  too,  the  composition  of  the  cells 
is  found  to  be  similar  indeed  to  that  of  the  parent  cells,  both  in  re- 
gard to  primary  and  secondarj^  constituents.  Thus,  Bang  found  that 
sarcomas  derived  from  lymph-glands  contain  the  particular  nucleo- 
proteins  that  are  found  normally  only  in  lymph-glands,  h^q^erne- 
phromas  contain  much  fat,  lecithin,  and  cholesterol :  squamous  cell 
carcinomas  develop  great  amounts  of  kerato-hyalin ;  carcinomas  of 
mucous  membranes  may  contain  much  mucin,  etc. 

Many  have  sought  in  cancer  tissues  a  poison  that  might  account  for 
the  cachexia  characteristic  of  new-growths.  Extracts  have  been  ob- 
tained that  were  destructive  to  red  corpuscles  (hemolytic),  and  that 
were  sometimes  slightly  toxic  to  animals,  but  the  results  have  not 
seemed  sufficiently  striking  to  account  for  the  appearance  of  cachexia. 
Because  of  the  interference  with  circulation,  brought  about  in  tumors 
by  pressure  of  the  growing  tissues  upon  their  blood-vessels,  areas  of 
necrosis  frequently  develop,  and  these,  undergoing  autolysis,  yield  sub- 
stances that  are  hemolytic  and  toxic.  "Whether  these  are  the  cause  of 
cancer  cachexia,  however,  may  be  questioned ;  but  they  are  sufficient  to 
account  for  most  of  the  experimental  results  as  yet  obtained.     No  sub- 

1  Literature  given  bv  Xeuberp,  Zeit.  Krel)sforscli..  1910  (10).  55;  and  Bhuiion- 
thal,  Ergebnisse  Physiol.,  1910   (10),  3(5.3. 

2  See  Wells  and  Long,  Zeit.  Krebsforsch.,  1013   (12),  598. 

492 


THE  CUE  Ml  ST  UY  OF  TUMORfi  493 

stance  has  yet.  been  isolated  from  or  detected  in  malignant  growths 
that  is  peculiai-  to  lliem  and  not  found  in  normal  cells,  and  still  less 
has  any  substance  been  detected  that  accounts  in  any  way  either 
for  the  occurrence  of  tumors  or  for  tlie  effects  that  they  produce. 

Tumor  cells  seem  to  depend  upon  much  the  same  conditions  as  nor- 
mal body  cells  for  their  growth,  since  anything  that  leads  to  wasting, 
malnutrition,  or  atrophy  in  the  tissues  of  the  host  usually  tends  to 
impede  the  rate  of  growth  of  the  tumor  cells,  in  marked  contrast  to 
infectious  diseases.  Specific  attempts  to  modify  tumor  growths  by 
diets  (Mendel-Osborne  diet)  which  stunt  the  animals  because  lack- 
ing certain  amino-acids  necessary  for  growth,  have  been  successful,-'' 
but  it  is  dilBcult  to  be  sure  that  this  effect  depends  on  the  specific  ab- 
sence of  a  definite  substance  rather  than  on  general  malnutrition.-*' 
Tumor  cells  made  incapable  of  utilizing  carbohydrate  through  com- 
plete phlorizin  diabetes  -'^  may  be  unable  to  grow,  and  even  retro- 
gress completely.  Furthermore,  the  constituents  of  the  hypophysis 
that  stimulate  somatic  tissue  growth  also  stimulate  growth  of  tumor 
tissues.-'' 

The  discovery  by  B.  Fischer^  that  fat  stained  with  scarlet-R  and 
injected  beneath  the  skin  causes  epithelial  proliferation  resembling 
but  not  terminating  in  cancer,  has  led  to  much  speculation  as  to  the 
nature  of  substances  which  might  cause  cells  to  proliferate  lawlessly 
and  malignantly.  The  great  frequency  of  cancer  in  workers  in  prod- 
ucts of  destructive  distillation  of  wood  (tar,  soot,  paraffin  *)  has 
also  indicated  the  possibility  of  chemical  stimuli  causing  cancers.  A 
striking  instance  of  chemical  stimulation  causing  cancer  formation 
is  furnished  by  the  cases  of  carcinoma  of  the  urinary  bladder,  which 
is  a  common  cause  of  death  in  men  who  work  in  aniline  dyes,  both 
dyers  and  dye  makers  being  subject  to  this  condition.  The  dyes  that 
seem  to  be  responsible  belong  to  the  group  of  aromatic  amido-hy- 
droxyls,  including  safranin,  congo-red,  benzopurpurin,  fuchsin,  eosin 
and  others.'^  H.  C.  Ross "  has  made  extensive  studies  of  the  relation 
to  cancer  of  substances  which  cause  leucocytes  to  multiply,  designat- 
ing them  as  "auxetics."  These  seem  to  be  present  in  the  anthracene 
fractions  of  tar,"""  which  may  explain  the  frequency  of  cancer  in  work- 
ers in  tar,  soot  and  paraffin.  Japanese  investigators  report  that  pro- 
tracted irritation  of  rabbits'  ears  with  tar  leads  to  strikingly  infiltra- 

2a  See  Sweet,  et  a1.,  Jour.  Biol.  Cliem.,  191.5    (21),  .300. 

2b -Rous.  .Tour.  Exp.  Med.,  1914    (20),  4.3.3. 

2c  Benedict  and  Lewis,  Proc.  Soe.  Exp.  Biol..  1914   (11)     134. 

2d  Robertson  and  Burnett,  Jour.  Exp.  Mod.,  1916   (23),  631. 

3  Verb.  Dent.  Path.  HeselL,  1906  (10).  20;  see  also  TTasra.  Zcit.  Krehsforscli.. 
1913  (12),  .52.5:  Saelis,  Wien.  klin.  Wocli.,  1911  (24),  1.5r)l:  Stoe])pr,  Muncli. 
nied.  Woch.,  1910   (57).  7.39  and  947. 

•tSee  Bavon,  Laneet.  1912    (ii),  1579. 
-      5  See  Leiienberffer,  Beitr.  klin.  Chir..   1912    (SO),  208, 

6  "Researches  into  Induced  Cell  Reproduction  and  Cancer,"  London. 

eaNorris,  Biocliem.  Jour.,   1914    (8),  253. 


494  THE    CHEMISTRY    OF    TUMORS 

tive  proliferation  of  the  epitlieliuni,  but  witliout  metastasis,  and  re- 
trogressing: when  the  irritant  is  discontinued.'"'  The  influence  of  vari- 
ous salts  on  cell  o;ro\vth  has  also  been  a])plied  to  cancer  patholofjy,  and 
while  we  have  abundant  evidence  that  chemical  substances  may  either 
stimulate  or  check  cell  growth,  as  well  as  regulate  it,  our  biological 
chemistry  has  not  yet  given  us  any  very  substantial  facts  on  these 
problems/ 

Nevertheless,  numerous  observations  have  been  made  concerning  the 
chemistry  of  tumors,  which,  although  they  do  not  as  yet  throw  any 
important  light  on  the  fundamental  problems  of  tumor  pathology, 
are  of  much  interest.     These  may  be  briefly  summarized  as  follows : 

A.     CHEMISTRY  OF  TUMORS  IN  GENERAL 

(1)  Proteins. — Earlier  studies  showed  that  tumor  growths  contain 
the  same  sorts  of  proteins  as  do  normal  tissues,  and  apparently  in 
about  the  same  proportions,  and  in  spite  of  certain  contradictory  re- 
ports this  statement  seems  to  be  correct. 

In  all  probability  the  nucleoproteins  of  tumors  share  the  specific 
characteristics  of  the  nucleoproteins  of  the  tissues  from  which  they 
arise — at  least  this  is  the  case  with  the  nucleoproteins  of  lymphosar- 
coma, according  to  Bang.-  The  characteristic  constituent  of  lymph- 
glands,  spleen  and  thymus  is  a  compound  of  nucleic  acid  and  histon 
{Jiistmi  nudeinate) .  If  to  a  watery  extract  of  an  organ  a  few  drops 
of  CaCL  solution  are  added,  the  formation  of  a  precipitate  indicates 
the  presence  of  a  lymphatic  tissue.  If  this  precipitate  is  soluble  in  1 
per  cent.  NaCl,  it  is  a  nucleinate  corresponding  in  type  to  that  of  the 
lymph-glands  and  spleen ;  if  not  soluble,  it  is  of  the  type  of  the 
thymus  or  leucocytes.  Extracts  from  no  other  organs  give  a  pre- 
cipitate with  calcium  chloride.  Spindle-cell  sarcomas  were  found  not 
to  give  this  reaction,  but  round-cell  sarcomas  of  lymphatic  origin  do, 
for  they  contain  the  specific  nucleinate  abundantly.  Bang  believes 
that  this  reaction  can  be  used  to  distinguish  sarcoma  arising  from 
lymphoid  tissue.  This  seems  to  have  been  confirmed  by  Bcebe,"  who 
found  nucleo-histon  only  in  lymph-gland  tissue,  but  the  distinction  be- 
tween thymus  and  lymph-gland  nucleohistou  is  probably  not  so  easily 
made  as  Bang  intimates. 

Because  of  their  richly  cellular  structure  cancers  may  contain  move 
nucleoprotein  than  the  tissues  from  which  they  arise.  Thus  Petry  ^^ 
found  50  per  cent,  of  nucleoprotein  in  carcinoma  of  the  mannuary 
gland,  as  against  30  per  cent,  in  normal  tissue,  which  is  perhaps  ve- 

«b;Mitt.  Med.  Oosollsch.,  Tokio,  1910    (im) .   1. 

7  A  thoory  of  foil  division   in  cancor  as  a   icsuK    of  (.'kH'tric   forces  is  uivon  by 
JoRsup  et  ah,  Tlioc-licm.  Jour.,  1909    (J),  19]. 
«  TTofinoistcr's  Boitr.,   199.3    (4),  36H. 
t>  Anipr.  Jour.  Phvsiol.,  190r)    (1.3),  341. 
loZeit.  physiol.  Chem.,  1899   (27),  398. 


CHE  Ml  ST  in    or   TUMONS  J\   (IKSEK.XL  495 

lated  to  Bergell's  iiiuliii<i-  of  a  liiyh  diunujio-acid  coiitont  in  mouse 
cancer.^^  However,  AVells  and  Long '-  found  the  proportion  of  purine 
nitrogen  in  tumors  of  several  classes  to  be  much  lower  tlian  might 
be  expected  from  the  nuclear  content  as  shown  by  the  microscope; 
also  Satta/^  found  unexpectedly  low  phosphorus  figures  and  Yosh- 
imoto/*  found  no  parallelism  between  the  number  of  nuclei  and  the 
nuclein  content.  The  purines  present  in  tumor  tissues  are  quite  the 
same  in  nature  and  proi^ortion  as  in  normal  tissues  (Wells  and  Long), 
as  also  are  the  nucleoproteins. 

Bergell  and  Dorpinghaus  ^■'  have  studied  the  nature  of  the  proteins 
in  tumors  by  determining  the  proportion  of  the  various  amino-acids 
that  compose  them.  Because  of  the  amount  of  material  necessary 
for  the  ester  method,  they  were  obliged  to  use  a  mixture  of  various 
primary  and  secondary  cancers  and  one  sarcoma.  The  protein  of 
this  tumor-mixture  was  characterized  by  the  very  high  proportion  of 
alanine,  glutaminic  acid,  phenylalanine,  and  asparaginic  acid,  there 
being  from  5  to  10  per  cent,  of  each.  Leucine  was  very  low,  5-10  per 
cent.,  as  against  20  per  cent.,  or  higher,  found  in  most  normal  tissues. 
Glycocoll  and  tyrosine  were  present  in  small  quantities,  and  serine 
was  probably  also  present.  Neuberg  '^^'  found  in  cancer  protein  1.3 
per  cent,  of  tyrosine,  17  per  cent,  of  leucine,  scarcely  1  per  cent,  of 
■glutaminic  acid,  and  4.92  per  cent,  of  glycocoll.  In  five  human  tu- 
mors of  different  sorts,  Kocher  ^^^  found  very  high  figures  for  dia- 
mino-nitrogen,  which  he  correlates  with  the  growth  function  of  lysine ; 
his  averages  were :  arginine,  12.42 ;  histidine.  4.86 ;  lysine,  11.23 ; 
total,  28.47  per  cent,  of  the  protein  nitrogen.  Strange,  and  as  yet  un- 
explained, variations  in  tryptophane  content  in  various  tumors  were 
found  by  Fasal,^*"'  some  having  a  very  high  tryptophane  figure,  while 
in  others  none  could  be  found.  Centanni  ^'"'  found  that  tiyptophane 
and  tyrosine  inhibit,  while  skatole  and  indole  stinuilate  carcinoma 
growth. 

Certain  authors  have  believed  that  the  cancer  cell  has  a  specific 
chemistry,^"  but  most  of  these  analyses,  including  that  of  Abderhalden 
and  ^Fedigreceanu,^*  seem  to  indicate  that  cancer  proteins  have  much 
the  same  composition  as  normal  proteins.  Cramer  and  Pringle  ^°  find 
that  there  is  less  nitrogen  in  mouse  cancers  than  in  equal  amounts  of 
other  mouse  tissue,   the  decrease  being   in   the   coagulable   nitrogen, 

11  Zeit.  f.  Krebsforsch.,   1907    (5),  204. 

12  ZbifZ.,  1913    (12),  .598. 

13  Arch.  Ital.  Biol.,   1908    (49),  .380. 
i-t  Biochem.  Zeit.,  1909    (22),  299. 
i-'Deut.  mod.  Woch.,   190.5    (31),   1420. 

i«  Arl).  a.  d.  Path.  Inst,  zii  Berlin,  190G,  p.  .593. 

i«a,Tour.  Biol.  Chem.,  1915   (22),  295. 

i''b  Biochem.  Zeit.,  1913    (55),  88. 

I'"- Tumori,  1913    (2). 

1- Blunienthal,  Zeit.  Krehsforseh..   1907    (5),   183. 

IS  Zeit.  phvsiol.  Chem.,  1910    (09),  60. 

laProc.  Royal  Soc.,  B.,  1910  (82),  315;  Jour.  Phvsiol.,  1910   (50),  322. 


496  THE    CHEMISTRY    OF    TUMORS 

incoaon^ilable  nitrogen  being  relatively  increased;  a  given  amount  of 
nitrogen  produces  more  cancer  than  normal  tissue.  The  water  con- 
tent of  rapidly  growing  tissues,  whether  normal  or  cancerous,  was 
found  to  be  high.  This  corresponds  with  the  analysis  of  Robin,-" 
who  found  the  water  content  high  and  nitrogen  low  in  carcinomas  of 
the  liver,  sulphur  being  especially  low,  and  Chisholm  -^  has  found  the 
proportion  of  nitrogen  in  several  human  tumors  lower  than  in  the 
somatic  tissues.  However,  the  lack  of  any  marked  specific  individual- 
ity of  cancer  proteins  when  tested  by  imnnmological  reactions,  indi- 
cates a  very  close  chemical  agreement  with  normal  tissue  proteins. 

On  account  of  the  amount  of  autolysis  going  on  in  tumors  the 
products  of  protein  splitting  are  usually  present.  Beebe  --  found  in 
a  number  of  tumors  leucine,  tyrosine,  tryptophane,  proteoses  (biuret 
reaction),  and  in  one  glycocoll.  Because  of  the  deficient  circulation  in 
the  tumors,  the  amino-acids  accumulate  in  the  cancer  tissues  in  suffi- 
cient amounts  to  be  detected,  and  may  be  found  even  when  no  macro- 
scopic evidences  of  degeneration  are  present.  Possibly  om  account  of 
this  poor  absorption  no  proteoses,  peptones,  or  amino-acids  could  be 
found  in  the  urine  of  cancer  patients  by  AYolff :  -^  but  T^ry  and 
Lilienthal  -*  found  a  positive  reaction  for  albumose  in  the  urine  in 
about  two-thirds  of  all  carcinoma  "cases  examined  by  them;  however,  it 
may  be  absent  even  in  advanced  stages.  Lactic  acid  is  also  present 
in  tumors,  according  to  Fulci  -®  and  Saiki,-^  the  latter  finding  0.-18 
gm.  of  lactic  acid  per  100  gms.  cancer  of  the  stomach. 

(2)  Other  Organic  Constituents. — These,  in  general,  resemble 
the  organic  constituents  of  the  tissue  from  which  the  tumor  arises,  for 
a  structural  resemblance  to  the  parent  tissue  always  exists,  and  as 
structural  features  depend  largely  on  the  proportion  of  the  chemical 
components,  a  structural  similarity  fairly  implies  a  chemical  simi- 
larity. For  example,  adrenal  and  renal  tissue  contain  much  lecithin 
and  cholesterol,  and  hypernephromas  show  a  similar  composition;  the 
fat  of  a  lipoma  is,  in  its  qualitative  features,  almost  identical  with  the 
normal  fat  of  the  same  individual ;  tumor  melanin  shows  no  charac- 
teristic chemical  distinction  from  normal  melanin,  etc. 

Glycogen  lias  been  particularly  studied  in  tumors,  especially  be- 
cause of  the  erroneous  idea  advanced  by  Brault  that  the  quantity  of 
glycogen  is  in  direct  proportion  to  the  malignancy.  From  a  summary 
of  all  the  evidence,  it  seems  that  two  chief  factors  determine  the  pres- 
ence and  amount  of  glycogen  in  tumors.  One  is  tlie  embryonic  origin 
of    the    tumors;    thus    tumors    of    cartilage,    striated    muscle,    or    of 

20  Cent.  Plivs.  Path.  StoflFwechs.,  1011    fd),  .'i77. 

21  Jour.  Pa'tliol.  and  l^act.,  1013    (17).  G06. 
22Ainer.  Jour.  Physiol.,  1004    (11),  130. 

23  Zeit.  f.  Krclisforschuiip,  1005    (3),  0,5. 

24  Arch.  f.  V(T(laiuiii-:skr.,  100.5    (11),  72. 
21  Gaz.  interna/.,  di  nied.,  1010,  No.  24. 

27  Arch.  mC'd.  e\[>6r.,  1011    (23),  370. 


viii:mistIx'Y  of  tumohs  j\  aESEitAi,  497 

squamous  epitlieliuni,  whiuli  tissues  iioniuilly  cuntaiu  inucli  g-lyeogen, 
are  likewise  provided  with  an  abundance  of  this  material.  Second, 
the  occurrence  of  areas  of  impaired  cell-iuitrition  favors  the  accumu- 
lation of  lilycogen  in  the  degenerating  tumor-cells,  just  as  it  leads  to 
a  similar  accumulation  in  all  other  tissues  (Gierke).-^  The  most  ex- 
tensive consideration  of  this  topic  is  reported  by  Lubarsch,-"  who 
found  glycogen  microscopically  in  447  (or  29  per  cent.)  of  1544  tumors 
examined.  Jt  was  i)resent  in  but  3  out  of  184  fibromas,  osteomas, 
glionuis,  hemangiomas,  lipomas,  and  lymphangiomas,  and  in  but  2  out 
of  260  adenomas  from  various  parts  of  the  body.  It  occurred  in  all 
teratomas,  rhabdomyomas,  hypernephromas,  and  syncytiomas.  In  138 
sarcouuis  glycogen  was  present  in  70  (50.7  per  cent.)  ;  of  415  carcino- 
mas it  was  found  in  181  (43.6  per  cent.).  In  the  squamous  epithelial 
cancers  70  per  cent,  contained  glycogen,  while  the  mucoid  or  colloid 
cancers  were  always  free  from  glycogen.  The  glycogen  undoubtedly 
enters  the  cells  from  without,  probably  entering  as  sugar,  and  being 
converted  into  glj'cogen  by  intracellular  enzymes.  "We  have  no  re- 
liable studies  of  the  actual  quantity  of  glj'cogen  in  various  tumors,  al- 
though Meillere  ^°  states  that  the  microscopic  and  chemical  examina- 
tion of  tumors  give  corresponding  comparative  results,  which  Gierke 
states  is  generally  true  with  glycogen  estimations. 

Pentoses. — Neuberg  ^^  reports  finding,  as  a  product  of  autolysis  of  a 
carcinoma  of  the  liver,  a  pentose  which  was  not  produced  by  autolysis 
of  either  normal  liver  tissue  or  the  primary  growth  in  the  stomach. 
Beebe  ^-  found  that  in  carcinoma  of  the  mammary  gland  the  per- 
centage of  pentose  {xylose)  is  somewhat  higher  than  the  amount  in 
normal  mammary  glands  (about  0.23  per  cent.).  Carcinoma  in  the 
liver  did  not  show  any  constant  excess  of  pentose  above  that  of  normal 
liver  tissue  (about  0.38  per  cent.).  A  primary  carcinoma  of  the  liver 
showed  quite  the  same  pentose  and  phosphorus  content  as  normal 
liver  tissue.  In  general,  no  constant  relation  of  pentose  to  origin, 
malignancy,  or  degeneration  of  tumors  was  observed. 

Purines  and  Purine  Enzymes. — The  purines  of  both  benign  and 
malignant  tumors  have  been  studied  by  Wells  and  Long,^"  who  found 
them  the  same  as  those  in  nonnal  tissues,  and  in  much  the  same  rela- 
tive proportions.  The  proportion  of  the  total  nitrogen  of  tumors  which 
is  constituted  by  the  purine  nitrogen  is  less  than  would  be  expected 
from  the  histological  evidence  of  the  amount  of  nuclear  material  con- 
tained in  the  tumors.  Tumors  also  seem  to  contain  much  the  same 
purine  enzymes  as  the  normal  tissues.  Thus,  guanase  seems  uni- 
versally present  in  tumors  derived  from  human  tissues,  and  adenase 

28  Ziegler's  Beitr.,  1005    (37),  502. 
29Virchow's  Arch.,  1006   (183),  18S. 
3oCompt.  Pvend.  Soc.  Biol.,  1000   (52),  324. 
31  Berl.  klin.  Woch..  1004   (41).  lOSl ;    1005    (42),  118. 
32Amer.  Jour.  Physiol.,  1005    (14),  231. 
33  Zeit.  f.  Krebsforsch.,  1013    (12),  508. 
32 


498  THE    CHEMIi^TIiV    OF    TUMORS 

is  missing,  although  autolyzing  tumors  can  disintegrate  their  nucleic 
acid  (nuclease)  and  change  the  adenine  radicals  of  the  nucleic  acid 
into  hypoxanthine,  presumably  by  way  of  adenosine  and  inosine 
(Amberg  and  Jones).  Secondary  tumors  growing  in  the  human  liver 
do  not  accjuire  the  enzyme,  xanthine-oxidase,  which  is  a  characteristic 
enzyme  of  tliis  organ.  The  liver  tissue  between  the  cancer  nodules 
seems  to  oxidize  purines  less  activelj^  than  normal  liver  tissue.  Long  ^* 
has  also  found  similar  conditions  in  tumors  from  sheep,  pigs  and 
cattle,  observing  that  primary  carcinoma  of  the  liver  does  not  con- 
tain xanthine  oxidase,  a  point  of  interest  in  view  of  the  fact  that  in 
the  development  of  mammals  the  xanthine  oxidase  does  not  appear 
until  late. 

Lipins. — Tumor  cells  seem  to  contain  much  the  same  fats  and  lipoids 
as  normal  cells,  and,  so  far  as  known,  in  much  the  same  pro])ortions 
as  characterize  the  cells  from  which  the  tumors  arose.  Thus  Wells  ^^ 
found  that  hypernephromas  show  the  same  high  proportions  of  leeitlii]i 
and  cholesterol  as  he  found  in  normal  adrenal,  and  as  are  found  in 
the  renal  cortex.  Other  malignant  tumors  have  much  less  lipoids  and 
fats  (see  Hypernephromas).  A  secondaiy  carcinoma  of  liver  cells^ 
metastatic  in  the  skull,  was  found  by  Prym  ^^  to  show  the  same  sort 
of  fatty  infiltration  that  is  characteristic  of  fatty  liver  cells.  On 
account  of  the  poor  blood  supply  of  many  tumors,  fatty  changes 
are  usual,  occurring  under  the  same  conditions  and  showing  the  same 
microscopic  features  as  fatty  degeneration  in  other  tissues,^^  being 
more  common  in  malignant  than  in  benign  tumors ;  especially  abun- 
dant in  squamous  cell  carcinomas,  and  scanty  in  sarcomas.  Crystals 
of  cholesterol  or  cholesterol  compounds  are  described  in  tumors  by 
White.^*  Even  lipoma  fat  shows  no  difference  from  normal  fat,^** 
and  the  depot  fat  of  tumor  patients  is  quite  the  same  as  in  patients 
with  other  diseases  associated  with  equal  wasting,^'^  in  whom  some 
increase  in  unsaponifiable  material  (cholesterol)  is  usual.  INIurray '*^ 
saj's  that  the  lipoids  of  degenerating  uterine  fibroids  are  strongly 
hemolytic,  which  may  account  for  the  so-called  ''red  degeneration"  of 
these  tumors.  Freuncl  and  Kaminer  *^  suggest  that  the  fatty  acids  of 
tissues  are  of  importance  in  determining  whether  a  tissue  is  a  suitable 
soil  for  secondary  growth,  these  substances  being  deficient  in  tissues 
where  growths  develop.  There  has  been  some  effort  to  correlate  the 
cholesterol  and  lecithin  contents  of  blood  and  tissues  with  the  rate  of 

34, Tour.  Exper.  Med.,  1013    (18),  rA2. 
35  .Tour.  Med.  Res.,  1908   (17),  461. 
3«l'>ankf.  Zeit.  Palli.,   1012    (10),   170. 

37  See  lliifra,  IJorl.  klin.  Wocli.,  ]:;]2  (40),  ,342;  Joamiovics,  Wien.  klin.  Woeli., 
1912    (25).  37. 

38  .Tour.  Patli.  and   Haci..  1!)0S    (I.']).  .3. 

39  See  WellH,  Arcli.  Int.  .Med.,  1912   (10),  2!l7. 

■»"  \\'acker,  Zeit.  ])hyfiiol.  Cheni.,  1!)12    (7S),  :54!>;    l!)12    (SO),  ;5S.3. 
41  Jour.  Obst.  Gvn.'lJrit.  Knip.,  1010    (17),  534. 
4-:Wien.  klin.  Woch.,  1012    (25),  KiOS. 


JXOh'dAMC  COX^TITLEXTS  OF  TUMOUH  499 

cancer  o;rowth ;  apparently  lecithin  inhibits  gi-o\vth  and  cholesterol 
stimulates/''  However,  Bullock  and  Cramer  *^''  found  much  more 
cholesterol  in  a  slowly  jifrowing-  mouse  carcinoma  than  in  a  rapidly 
growing  one.  somewhat  more  phosphatid  in  the  latter,  much  more 
phosphatid  in  a  sarcoma  than  in  the  carcinoma,  and*  cerebrosides  only 
in  the  latter;  in  necrotic  portions  of  tumors  they  found  an  increase  in 
simple  fats.  These  tigiires  are  based  on  too  few  observations  to  be  in- 
terpreted as  yet. 

(3)  Inorganic  Constituents. — These  have  been  studied  under  ex- 
ceptionally favorable  conditions,  in  that  the  age  of  the  tumor  could  be 
accurately  estimated,  in  the  inoculable  carcinoma  of  mice  (Jensen), 
by  Clowes  and  Frisbie.^^  They  found  that  rapidly  growing  tumors 
contain  a  high  percentage  of  potassium  and  little  or  no  calcium, 
whereas  in  old,  slowly  growing,  relativel}'  necrobiotic  tumors  the  rela- 
tion is  reversed,  the  potassium  decreasing  greatly  while  the  calcium  in- 
creases. Magnesium  is  present  only  in  traces,  while  the  proportion  of 
sodium  fluctuates  much  less,  but  is  usually  greater  than  either  the 
potassium  or  calcium,  although  in  very  old  tumors  the  latter  may  be- 
come excessive.  The  most  rapid  growth,  however,  seems  to  occur  in 
tumors  in  which  both  calcium  and  potassium  arfe  present  in  the  ratio  of 

K  2  3 
—  =  -  or  - 
Ca         1         2 

Beebe  ^''  analyzed  a  number  of  human  tumors  with  the  following 
results:  PhosphoTOS  was  found  in  proportion  to  the  amount  of  nu- 
clear material,  varying  from  0.139  per  cent,  (uterine  fibroid)  to  1.06 
per  cent,  (sarcoma).  Iron  varied  from  0.013  per  cent,  to  0.064  per 
cent.,  probably  depending  on  the  amount  of  blood  and  nucleoproteins. 
Calcium  is  most  abundant  in  old  degenerated  tumors,  and  potassium  in 
rapidly  growing  tumors.  These  results,  supported  by  Clowes  and 
Frisbie's  findings,  indicate  the  importance  of  potassium  for  cell 
growth.  Injection  of  potassium  salts  into  mice  increases  their  suscep- 
tibilitj'  to  inoculation  (Clowes),""^  while  calcium  decreases  cancer 
growth  (Goldzieher).'*"  A  greater  proportion  of  potassium  was  found 
in  primary  than  in  secondary  growths  by  Mottram ;  '*^  sodium  was  the 
same  in  each ;  there  is  more  potassium  in  squamous  cell  carcinoma 
than  in  round  cell  sarcoma.  Robin  ^^  states  that  in  cancerous  livers 
the  cancer  tissue  contains  more  inorganic  matter  than  the  normal  liver 

43  See  Robertson  and  Burnett.  Jour.   Exp.  Med.,   1013    (17),   344;    1916    (23), 
631;   Sweet  et  al..  Jour.  Biol.  Cliem.,  1915   (21),  309. 
43aProc,  Roval  Soc,  London    (B),  1914    (87),  236. 
4*Amer.  Joiir.  Physiol.,  190.5   (14),  173. 
45  Amer.  Jour.   Plivsiol.,   1904    (12),   167. 
48  British  Med.  Joiir.,  Deo.  1,   1906. 
4TVerhandI.  Deiit.  Path.  Gesellsch.,   1912    (15),  283. 
48  Arch.  Middlesex  Hospital,    1910    (19),  40. 
49Conipt.  Rend.  Acad.  Sci.,  1913   (156),  334. 


500  THE    CHEMISTRY    OF    TUMORS 

tissue  about  it.  Cattley  •""'  found  tiie  inierochemic  distribution  of  po- 
tassium the  same  in  cancer  as  in  normal  cells,  and  the  same  seems  to  be 
true  of  maup:anese.'''^ 

Schwalbe  ^^  found  that  cancer-cells  contain  iron  in  a  condition 
demonstrable  b}^  the  Berlin-blue  reaction,  and  occurring  independent 
of  hemorrhages.  Tracy  ■'*  found  that  tumors  reacted  microscopically 
for  iron,  either  free  or  in  the  form  of  an  albuminate,  only  in  areas 
where  hemorrhages  had  occurred.  Nuclear  or  organic  iron  could  be 
detected  in  the  nuclei,  occurring  in  a  network  arrangement.  In  other 
words,  iron  occurs  in  tumors,  both  quantitatively  and  qualitatively, 
exactly  as  in  normal  cells  of  the  same  type.  The  same  writer  ^^  found 
in  tumors,  by  microchemical  reactions,  that  phosphorus  in  the  form 
of  nucleo])roteins  likewise  shows  no  essential  dift'erences  from  its  dis- 
tribution in  normal  tissues. 

In  this  connection  may  be  mentioned  the  observations  of  Hem- 
meter,""  who  found  that  the  cells  of  carcinoma  of  the  mammarv 
gland  will  shrink  when  placed  in  physiological  salt  solution  or  in 
the  serum  of  the  patient,  whereas  normal  cells  swell  when  placed  in 
cancer-juice.  This  suggests  that  the  osmotic  pressure,  and,  by  infer- 
ence, the  amount  of  inorganic  constituents,  is  lower  than  in  normal 
tissues.     Crj'stalloids,  such  as  KI,  diffuse  readily  into  cancer  tissue. ^'•'^ 

(4)  Enzymes. — The  rapid  and  extensive  autolysis  that  occurs  in 
tumors,  as  showm  both  morphologically  and  by  the  presence  of  the 
products  of  protein  cleavage  in  them,  indicates  that  tumor  cells 
resemble  all  other  cells  in  possessing  intracellular  proteolytic  enzymes. 
Because  of  autolysis,  puncture  fluids  in  cancer  of  serous  surfaces  show 
an  increased  amount  of  incoagulable  nitrogen  (Morris), '''  and  they 
may  show  free  amino-acids  (Wiener)  j"^^  while  there  is  a  slight  increase 
in  the  incoagulable  nitrogen  of  the  blood  (Takemura)."'** 

There  is  considerable  but  not  undisputed  evidence  that  cancer  tis- 
sue autolyzes  somewhat  more  rapidly  than  corresponding  normal  tis- 
sues,**"  and,  according  to  Neuberg,  Blumenthal  and  others,'^^  that 
cancer  extracts  digest  other  tissues  than  themselves  (heterolysis),  a 
property  not  exhibited  by  extracts  of  normal  tissues.  ]\Iiiller  and 
others  would  ascribe  this  heterolysis  to  the  leucocytes  present  in  the 

soLancot,   1907    (172),  13. 

ci  Modifrrofoaini,  Proc.  Roval  Soc,  B,   1912    (86),  174. 
53  ('(.111.  f.   I'atli.,   inoi    (12),  874. 
f'^  Jour.  IMcd.  Kpscarch,  1905   (14),  1. 
55  Martha  Tracy,  Jour.  Mod.  rjescaroli,  1906    (14).  447. 
50  Auier.  Jour.  Med.  Soi.,  190.3   (125),  080. 

.Ida  Van  den  Volden,  P.ioohem  Zeit.,  1908  (9),  54;  aeo  also  Wells  ami  Tledon- 
hiiTfr,  Jour.  Infect.  Di.s.,  1912   (11),  349. 

57  Arch.  Int.  Med..   1911    (8),  457. 

58  Hiochem.  Zeit.,   1912    (41),  149. 
50  I  hid.,   1<)10    (25),  505. 

«oSc,.  ^(oliirnolo,  P»iochem.  Zeit.,  1909  (22),  299;  Daels  and  Dclenz^,  Bull. 
Acad.  .Med.  J'.clj;.,  1913    (26),  833. 

01  Bibliography  by  Hamburger,  Jour.  Amer.  Med.  Assoc,  1912  (59),  847. 


ENZYMES  OF  TUMORS  501 

tumors.  Nucleases  have  been  found  in  tumors  as  in  otlier  tissues,"^ 
and  in  general  the  enzymes  which  deamidize  adenine  and  guanine  (ad- 
enase  and  guanase)  are  usually  present  if  the  original  tissue  possessed 
these  enzymes,  but  no  instance  of  tlie  presence  of  xanthine  oxidase 
or  uricolytie  enzyme  has  been  obtained  (Wells  and  Long,  loc.  cit.^^). 

Hamburger  finds  that  the  enzymes  of  cancer  tissue  upon  which  the 
glycyl-tryptophane  and  other  enzyme  tests  for  cancer  are  based,  are 
ereptases,  resembling  in  all  thinr  properties  the  ereptases  of  -normal 
tissues,  and  not  })resent  in  particularly  large  amount.  However,  Ab- 
derhalden  ^^  has  found  evidence  that  certain  peptids  may  be  split  in  a 
different  way  by  cancer  than  by  normal  tissues,  supporting  those  who 
hold  that  cancer  enzymes  are  different  from  normal  tissue  enzymes. 
Autolysis  of  tumors  is  said  to  be  augmented  by  x-ray,  and  especially 
by  radium  (Neuberg),  and  tumor  tissue  is  readilj^  digested  by  tryp- 
sin. 

The  presence  of  ereptases  in  carcinomatous  gastric  juice  has  been 
especially  studied  because  of  its  diagnostic  possibilities,  and  the  care- 
ful investigation  of  Jacques  and  Woodyatt  "^  seems  to  show  conclu- 
sively that  such  an  enzyme  is  rarely  present  in  gastric  juice  except 
when  derived  from  a  cancer  present  in  the  wall  of  the  stomach,  pro- 
vided peptolytic  bacteria  are  excluded  by  filtration.  Deaminizing 
enzymes  may  also  be  found  in  gastric  cancer  secretions.*'^"  In  the  blood 
of  cancer  patients  there  is  usually  an  increased  antitrj^ptic  activity, 
ascribable  to  the  reaction  against  enzymes  absorbed  from  the  cancer; 
it  is  less  pronounced  with  sarcoma.*'^  The  body  tissues  of  patients 
dying  with  cancer  show  a  low  ereptic  activity,  but  the  same  is  true  of 
persons  d.ying  from  other  wasting  diseases  ( Colwell ) ."''  The  same 
seems  to  be  true  of  other  tissue  enzymes ; — at  least  purine  oxidizing 
enzymes  are  deficient  in  the  liver  tissue  between  secondary  cancers 
(Wells  and  Long^^)  and  the  eatalase  is  also  reduced  in  liver  tumors 
(Blumenthal  and  Brahn)  ^''  and  in  the  blood  of  tumor  mice  (Rosen- 
thal)''^ ;  in  human  blood  the  eatalase  may  vary  either  side  of  normal.®*^ 
Brahn  °^^  found  that  liver  metastases  of  gastric  cancer  contained  no 
lipase  or  lecithinase,  which  enzymes  were  also  reduced  in  the  liver  tis- 
sue between  cancer  nodules.  However,  choline  has  been  found  in 
necrotic  sarcomas  of  rats,"*''  which  would  seem  to  indicate  the  presence 
of  enzymes  disintegrating  lecithin.     As  mentioned  elsewhere   (Mela- 

62  Goodman.  Jour.  Exp.  Med.,  1912   (15),  477. 

63  Zeit.  Krebsforscli.,  1010    (9),  2fi6. 
6-tArch.  Int.  INled.,  1912  (10),  560. 

64aHalporn,  Mitt.  Grenz.  Med.  Chir.,  1915    (28),  709. 

65  Citronblatt,  Med.  Klin.,  1912   (8),  1.38. 

66  Arch.  Middlesex  Hosp.,  1909   (15),  96. 
6T  Zeit.  f.  Krebsforsoh.,  1910    (8),  436. 
csDeut.  med.  Wocli.,   1912    (.38),  2270. 

68a  Rohdonburo:,  X.  Y.  ]\Ied.  .Tour.,  1913    (97),  824. 
esbSitzber.  kpl.  preuss.  Akad.  Wiss..   1916    (20),  47-8. 
68c  Euinger,  Miinch.  mod.  Woch.,   1914    (61),  2336. 


502  THE    CHEMISTRY    OF    TUMORS 

nin),  melanotic  tumors  may  contain  enzymes  oxidizing  tyrosine,  epine- 
phrin,  pj^rocatechin,  or  other  related  aromatic  substances,  with  the 
formation  of  pigmentary  substances.  Catalase  is  low  in  tumor  tissues 
(Blumenthal  and  Brahn).     (See  also,  Autolysis  in  Tumors,  chap,  iii.) 

Other  enzymes  are  also  present  in  tumor  cells.  Buxton  '^^  exam- 
ined a  large  number  of  tumors  for  their  enzymes  by  the  plate  {aiixan- 
ographic)  method,  and  found  considerable  variations  in  different 
growths.  All  contained  amj'lase  (splitting  starch)  and  lipase  (split- 
ting butyrin).  IMost,  but  not  all,  tumors  coagulated  milk  and  liquefied 
casein,  and  also  liquefied  gelatin  (rennin,  proteases).  Peroxidase  was 
nearly  ahvays,  and  catalase  always,  present.  Digestion  of  fibrin,  co- 
agulated seinim,  and  coagulated  egg  albumen  could  not  be  observed. 
Practically  all  tumors  split  glycogen.  Tj-^rosinase  could  not  be  demon- 
strated. The  fact  that  early  embryonic  tissues  were  found  poor  in 
enzymes  ""^  speaks  against  the  common  assumption  that  tumors  repre- 
sent strictly  an  embryonic  formation,  but  Long  "^  found  that  xanthine- 
oxidase,  which  in  normal  development  does  not  appear  until  late  in 
fetal  life,  was  absent  from  primary-  carcinomas  of  sheep  livers,  al- 
though normal  adiilt  sheep  liver  tissue  is  rich  in  this  enzyme. 

]\lacFadyen  and  Harden  ^-  studied  the  juices  obtained  by  grinding 
up  tumor  cells  made  brittle  by  liquid  air,  and  found  by  direct  meth- 
ods (chiefly  in  breast  cancers)  invertase,  maltase,  amylase,  proteases 
acting  in  both  acid  and  alkaline  solutions,  catalase,  oxidase,  with  per- 
haps traces  of  lipase  and  peroxidase,  but  no  lactase. 

Tumors  arising  from  the  gastric  mucosa,  according  to  AYaring,'^^ 
contain  both  pepsin  and  rennin ;  those  from  the  pancreas,  both  pri- 
mary and  secondary  growths,  contain  trypsin,  steapsin,  amylase,  and 
rennin. 

(5)  Internal  Secretion. — If  tumors  are  derived  from  an  organ 
with  an  important  internal  secretion,  the  tumor  cells  in  many  cases 
produce  the  same  internal  secretion,  which  seems  to  have  the  same 
functional  properties  as  the  normally  produced'  secretion.  Thus  a 
metastatic  growth  from  a  thyroid  tumor  has  been  said  to  functionate 
in  place  of  the  resected  gland ;  Gierke  ^*  found  in  about  20  grams  of 
material  from  metastatic  thyroid  tissue  in  the  vertebral  column  about 
5  mg.  of  iodin,  which  was  a  trifle  larger  proportion  than  was  present 
in  the  thyroid  itself.  Carlson  and  Woelfel "''  found  much  iodin  in 
the  metastases  of  a  thyroid  carcinoma  of  a  dog,  while  in  another  dog 
whose  cancerous  thyroid  contained  no  iodin  the  secondary  tumors 
were  also  devoid  of  this  element.     ^Marine  and  Johnson  '^  found  that 

fin.Tour.  Med.  Research,   inn.3    (0),  :]r-,(\. 

■!0  Ihid.,  mOo   (l.*?),  .54.S. 

Ti.Toiir.  Expor.  Mod.,  101.3    (18),  512. 

-2  Lancet.    100.3    (ii),    224. 

73  .Tour.  .Anat.  and  Phvsiol.,  1804    (28).  142. 

74TTofrneisior's  ■Reitr.."l0n2    (3),  280. 

-".  Amer.  Jour.  Plivsiol..  1010   (20),  32. 

70  Arch.  Int.  Med."   1013    (11),  288. 


INTERNAL  SECRJJTIOX  OF  TUMORS  503 

in  two  cases  of  caucer  of  the  thyroid  in  man,  and  one  in  the  dog,  tlie 
cancer  tissne  sliowed  no  ability  to  retain  iodin  given  by  month,  in  con- 
trast to  nonnal  thyroid  and  simple  adenomas.  Meyer-Hiirlimann  and 
Oswald  '"'*  have  described  a  remarkable  ease  of  cystic  carcinoma  of  the 
thyroid,  from  which  in  six  weeks  2840  c.c.  of  secretion  was  obtained 
by  pnncture.  It  contained  0.077  mg.  iodin  per  10  c.c.  (the  patient 
having  previously  been  given  KI)  as  compared  with  normal  thyroid 
which  contains  0.4  to  4  mg.  per  10  gm.  It  contained  both  globulin  and 
albumin,  the  former  corresponding  to  true  thyroglobulin,  even  to  in- 
creasing vagus  irritability  experimentally.  The  "adenomatous" 
nodules  of  the  thyroid  often  show  evidence  of  active  secretion, 
Goetsch'*'''  having  found  their  cells  rich  in  mitochondria,  while  Gra- 
ham ^"'^  found  that  they  take  up  iodin  and  metabolize  it  so  that  the 
adenomatous  tissue  produces  the  typical  thyroid  effect  on  the  develop- 
ment of  tadpoles.  Adrenal  cancers  do  not  usually  cause  Addison's 
disease,  because  they  functionate  in  place  of  the  destroj^ed  gland 
(Lubarsch). 

In  the  peculiar  and  characteristic  production  of  cachexia,  often  ap- 
parently out  of  all  proportion  to  the  amount  of  tumor  tissue,  there 
would  seem  to  be  evidence  that  a  peculiar  and  abnormal  product  of 
metabolism  is  formed  by  cancer-cells,  and  extracts  from  cancers  have 
been  found  toxic  for  protozoa.^^  As  yet,  however,  it  has  been  im- 
possible to  demonstrate  any  characteristic  toxic  substance  in  cancers.'"'' 
Girard-^Iangin  '^  claims  that  malignant  tumors  contain  colloidal  poi- 
sonous substances  in  proportion  to  their  softness,  extracts  causing 
paralysis  and  fall  of  blood  pressure ;  but  others  have  failed  to  substan- 
tiate this.'-'  Because  of  the  constant  disintegration  of  the  tumor  tis- 
sues, products  of  autolysis  are  formed,  and  undoubtedly  enter  the  cir- 
culation in  small  quantities ;  possibly  they  are  a  factor  in  the  systemic 
manifestations  of  malignant  growths,  analogous  to  the  action  of  cleav- 
age products  of  foreign  proteins  which  may  produce  "protein  fever" 
and  other  toxic  effects. 

Since  all  normal  tissue-cells  produce  substances  through  their  me- 
tabolism that  enter  the  circulation,  it  is  quite  certain  that  tumor-cells 
do  likewise,  and  it  is  highly  probable  that  the  presence  of  abnormal 
quantities  of  such  products,  even  if  they  are  of  quite  normal  compo- 
sition, may  cause  disturbances  in  the  body.  As  yet,  however,  no  such 
substances,  either  normal  or  abnormal,  have  been  isolated,  nor  has 
their  presence  been  demonstrated.     Numerous  isolated  observations  of 

-6aKorr.-Bl.  Sohweizer  Aerzte,  1013   (43),  1468. 

76b  Bull.  Johns  Hopkins  Hosp.,  1916    (27),   120. 

76c  .Tour.  Exp.  :\rc(l.,  1916    (24),  34.>. 

T7  Woodruff  and  Underbill,  Jour.  Biol.  Cheni.,  1913  (15).  401;  Calkins,  .Tour. 
Cancer  Res..  1916    (1),  205  and  399. 

7  7a  See  Blunientlial,  Festschr.  f.  Salkowski,  Berlin,  1904;  HansenuTun,  Zeit. 
Krebsforscli.,   1906    (4),  565. 

TsPresse  Med..  1906,  p.  1709;  Compt.  Rend.  Soc.  Biol.,  1909   (67),  117. 

79  See  Brusehettini  and  Barloeco,  Cent.  f.  Bakt.,  1907    (43),  064. 


504  THE    CHEMISTRY    OF    TUMORS 

ptoinains  or  similar  substances  in  tlie  urine  of  cancer  patients  may  be 
found  in  the  literature,®"  but  their  importance  is  extremely  question- 
able. 

Hemolytic  Substances. — A  number  of  observers  have  described  the 
finding  of  hemolytic  substcinces  in  cancer  extracts.  Bard  ®^  observed 
that  in  hemorrhagic  carcinomatous  exudates  in  serous  cavities  the 
blood  is  rapidly  hemolyzed,  which  is  not  the  case  in  exudates  from 
other  causes,  but  this  was  not  corroborated  by  Weil.®'  Kullmann  ®*^ 
found  that  extracts  of  carcinomas  contain  hemolytic  substances  acting 
energetically  both  in  the  body  and  in  vitro;  these  are  soluble  in  alcohol 
and  in  water,  are  not  complex  in  composition,  are  not  specific  for  hu- 
man corpuscles,  but  are  toxic  for  all  varieties  of  corpuscles.  IMicheli 
and  Donati  ®^  likewise  found  hemolytic  substances  in  8  of  15  tumors, 
of  which  5  acted  on  all  varieties  of  corjDuscles,  and  3  acted  on  only 
certain  varieties;  they  regard  the  hemolj'tic  substances  as  the  products 
of  autolysis  in  the  tumors.  Weil  ®*  also  found  the  hemolytic  property 
of  tumor  extracts  to  vary  with  the  amount  of  necrosis,  from  which  are 
derived  dialysable  hemolytic  substances  distinct  from  the  hemolysins 
of  normal  tissues.  It  is  well  known  tliat  among  the  products  of 
autolysis  of  normal  tissues  are  hemolytic  substances.  Whether  the 
severe  anemia  frequently  present  in  carcinoma  is  due,  either  largely 
or  in  part,  to  these  products  of  autolysis  is  unknown,  but  it  is  very 
probable  that  they  have  some  effect. 

Hemolysis  in  Cancer. — The  blood  serum  of  cancer  patients  has  often 
a  hemolytic  action  on  the  corpuscles  of  normal  persons  (Crile),  but 
this  property  is  quite  inconstant,  being  present  in  67  per  cent,  of  a 
series  of  472  cancer  cases  collected  by  Krida,  while  15  per  cent,  of 
cases  of  other  diseases  and  2.6  per  cent,  of  normal  persons  showed 
hemolytic  activity  of  the  serum. ®^  Elsberg  found  that  normal  corpus- 
cles injected  subcutaneously  into  cancer  patients  are  hemolyzed,  but 
Gorham  and  Lisser  found  this  reaction  positive  in  but  60  per  cent,  of 
their  eases,  the  subcutaneous  hemolysis  not  corresponding  at  all  to 
the  hemolytic  activity  of  the  patient's  serum  in  the  test  tube.  The 
stomach  contents  in  cancer  of  the  stomach,  when  ulcerated,  are  hemo- 
lytic (Grafe  and  Rohmer).®*^  The  red  corpuscles  of  cancer  patients  are 
said  to  have  usually  a  greater  resistance  to  hemolysis  by  cobra- venom 
than  normal  corpuscles,  but  this  is  not  eluiracteristic,  tliere  being  simi- 
lar alterations  in  other  diseases.®'     The  reputed  power  of  the  serum 

80  See  Kullmann,  Zcit.  klin.  IMcd..   in04    (.')3),  2fl-'?. 

81  La  Scmaine  ^tecL,  1001    (21),  201. 

82  Jour.  ]\Iecl.  EcH..  1010   (23),  80. 

83  Rift.rma  Mod.,  100.1   (19),  10.37. 

84  Jour.  Mod.   Pu's.,   1007    (K!),  287. 

sn  Liioraturo  l»v  (iorliain  and  Liasor,  Amor.  Jour.  ^lod.  Si'i.,  li*]2    (144),  10.'?. 
80  Dcut.  Arcli.  klin.  ]\lod.,   1008    (04),  230. 

87  Kraua,  Kanzi  and  II.  Klirlidi,  Sit/..  Bor.  Akad.  Wion.,  1010  (110).  3:  spo  also 
Grunbaum,  Jour.  rath,  and  lUiot.,  1012   (17),  82. 


METMiOLlSM  i\  qAycEii  505 

in  eaiK'or  to  i)r()toc't  I'orjju.st-lc.s  fi-om  liemolysis  b}-  oleic  and  lactic  acid 
could  not  be  demonstrated  by  Sweek  aijd  Fleisher.'*^ 

An  extensive  review  of  the  literature  and  methods  led  Cohnreich  **" 
to  the  conclusion  that  resistance  of  erythrocytes  to  hypotonic  solutions 
and  to  poisons  vary  independently  of  one  another.  He  has  devised  an 
improved  method  for  testing  resistance  to  hypotonic  solutions,  which 
seems  to  vary  directly  with  the' amount  of  stroma  and  PO4  content, 
and  finds  that  determinations  of  maximum  and  minimum  resistance  is 
of  little  value,  as  these  concern  only  a  small  part  of  the  corpuscles; 
he  therefore  determines  the  "plurinnnn"  resistance,  involving  most  of 
the  corpuscles.  The  most  significant  results  were  obtained  in  cancer 
of  the  alimentary  tract,  in  which  an  increased  resistance  was  always 
demonstrable.  Farmachidis  ^^^  finds  the  cobra  venom  resistance  more 
specific  for  cancer  than  most  other  investigators. 

(6)  Metabolism  in  Cancer. — Speaking  against  any  specific  na- 
ture in  the  cause  of  cancer  cachexia  are  numerous  observations,  indi- 
cating that  the  cachexia  is  in  no  way  different  from  the  cachexia  of 
other  conditions.  The  behavior  of  the  nitrogen  metabolism  seems  to 
be  quite  the  same  as  in  tuberculosis  and  other  wasting  diseases.  There 
is  the  same  excessive  elimination  of  aromatic  substances  (phenol,  indi- 
can)  and  oxyacids  (Lewin,^®  Blumenthal  ^°),  which  Lewin  considers  to 
arise  from  the  abnormal  metabolism  of  proteins,  and  not  from  putre- 
factive decomposition  in  the  tumor  or  in  the  intestines.  In  rats  with 
sarcoma,  increased  excretion  of  uric  acid  and  creatin  has  been  ob- 
served.°°^  There  is  also  the  same  excessive  elimination  of  mineral 
salts  that  is  observed  in  pulmonarv^  tuberculosis,  and  termed  "demin- 
eralization"  by  Robin, ^^  but  no  alteration  in  the  excretion  of  chlo- 
rides."^'^ As  in  other  cachexias,  the  creatin  content  of  the  muscles 
is  decreased.''-  Fraenkel  ^^  finds  evidence  that  there  maj'  be  some  diffi- 
culty in  tryptophane  metabolism  in  tumors  and  in  tumor  patients. 
Extensive  respiratory  studies  by  "Wallersteiner  ^^'^  showed  enormous 
variations  in  the  amount  of  heat  production  in  different  cases,  in  about 
10  per  cent,  of  which  figures  as  high  as  those  of  severe  fevers  or 
exophthalmic  goiter  were  obtained  repeatedly;  most  of  the  cases 
showed  high  normal  figures.     Nitrogen  loss  did  not  ordinarilj^  occur  if 

ssJour.  Med.  Res.,  1013    (27).  3S.3. 

88a  Folia  HematoL,  1913   (16).  307.  full  bibliography. 

88b  Gaz.  degli  Osped.,   1915    (30),  fiSO. 

s9Deut.  med.  Woch.,  190,5    (31),  218. 

90  Festschr.  f.  Salkowski,  Berlin.  1904. 
90aOrdway,  Jour.  Med.  Res.,  1913   (23),  301. 

91  Quoted  by  Lewin,  loe.  cit.  Clowes  ct  al.  (5th  Ann.  Rep.,  X.  Y.  State  Dept. 
of  Health.  190.3-4)  report  observing  a  slight  chloride  retention  in  cancer  pa- 
tients, and  review  tlie  literature  of  metabolism  in  cancer. 

9ia  Robin.  Compt.  Rend.  Acad.  Sci..  1913    (156),  1262. 

92  Chisholm,  Riocliem.  Jour.,  1912    (6),  243. 

93  Wien.  klin.  Woch.,  1912   (25),  1041. 
93aDeut.  Arch.  klin.  Med.,  1914  (116),  145. 


506  THE  en  KM  I  ST  If  y  of  timors 

the  ealorimetric  finding's  were  considered  in  the  calculations;  nitrogen 
equilibrium  was  maintained  if  sufficient  nourishment  was  obtained 
and  utilized.  In  general,  metabolism  iu  cancer  resembles  that  of 
fever,  and  warrants  the  assumption  of  a  toxic  stimulation  of  tissue 
destruction.  It  is  entirely  possible  that  the  products  of  cancer  protein 
destruction  are  responsible  for  this  toxicogenic  metabolic  abnormality, 
since  Vaughan  has  demonstrated  that  the  effects  of  bacteria  and 
foreign  proteins  are  quite  the  same  in  their  pyretic  and  toxic  action. 

Salkowski  demonstrated  that  the  amount  of  colloidal  nitrogenous 
material,  precipitated  from  the  urine  by  strong  alcohol,  is  increased  in 
cancer.  Numerous  observers  have  corroborated  this,  but  find  that  a 
similar  condition  obtains  in  other  cachectic  diseases,  although  in  cancer 
the  amount  of  colloidal  nitrogen  seldom  is  as  low  as  normal  unless  the 
tmnor  is  removed.''*  ]\Iuch  of  this  colloidal  nitrogen  seems  to  be  in  the 
form  of  "oxy-proteic  acid"  (Salomen  and  Saxl),^^  which  is  a  mixture 
of  incompletely  oxidized  polypeptids,  containing  much  unoxidized  sul- 
phur. The  proportion  of  neutral  sulphur  in  the  total  sulphur  in  the 
urine  seems  to  be  increased  in  cancer  (Weiss),  but  not  so  constantly 
or  characteristically  as  to  be  of  diagnostic  value.^^  Much  clinical  in- 
vestigation has  been  made  of  these  urinary  changes,  which  has  gen- 
erally substantiated  the  fact  that  there  usually  is  more  increase  in 
colloidal  nitrogen  and  ethereal  sulphate  in  the  urine  of  cancer  than 
in  other  diseases,  but  that  in  no  sense  are  these  changes  specific  for 
cancer,  and  the  fundamental  metabolic  disturbances  responsible  have 
not  been  ascertained.^®^  They  seem  more  indicative  of  the  excessive 
catabolism  of  cachexia  than  of  cancer  tissue  itself.  Saxl  ^^^  has 
ascribed  part  of  the  increased  sulphur  elimination  to  abnormal  excre- 
tion of  sulphoeyanid,  and  as  small  doses  of  sulphocyanides  lead  to 
increased  oxyproteic  acid  in  the  urine  he  suggests  that  in  cancer  there 
is  a  specific  disturbance  in  sulphoeyanid  metabolism,  an  hypothesis 
that  awaits  confirmation.  Of  similar  status  is  the  excessive  excretion 
of  glycuronic  acid  described  by  Roger. ""- 

Israel,  and  also  Engelmann,  have  reported  the  occurrence  of  a 
marked  increase  in  the  lowering  of  the  freezing-point  of  the  blood  in 
carcinoma  (as  low  as  — 0.60°  to  — 0.63°,  the  normal  being  — 0.56°), 
which  they  attributed  to  the  presence  of  excessive  products  of  protein 
decomposition  in  the  blood.  Engel,^"  however,  found  no  such  increased 
lowering  of  the  freezing-point  in  his  cases,  and  questions  the  signifi- 

!>4See  Mancini.  Dent.  Arch.  klin.  l\Tod,.  I'Ml  (103),  288-.  Seniciuiw.  Folia  T'rol., 
1912    (7),  21.5;  de  Bloemo  et  nl.,  Bioolioin.  Zcit..  1014    (0.5).  .34;"). 

»•'••  WicTi.  klin.  Woch.,  1!)11    (24),  440. 

!"!  RliuUmiillor  and  Koscnblooni.  .\rcli.  Inf.  Alrd..  101:i  (12),  27{>;  Intorstato 
Med.  Jour.,  1916    (23),  Xo.  2:  bildiojrrapliy. 

f'Oa  See  Goodridfie  and  Kaliii.  Hioclicni.  l^iill..  10].")  (4).  US:  Damask.  ^Vion. 
klin.  Woch.,  lOl.')    (28).  499;   Rassa,  Biodicni.  Zoit..  1914    ( f)4 ) ,  19."). 

n«!b  Biodiom.  Zoit.,  1913   (.'5")).  224. 

oi'  P.ull.  Soo.  :Mpd.  Hop..  Paris,  191")    (31).  499. 

t'TBcrl.  klin.  Woch.,  1904   (41).  82S. 


IMMUNITY  RfJACTlOXS  IX  CANCER  507 

cance  of  the  results  of  Israel  aiul  Eiigelniann.  According  to  Moore 
and  Wilson  "**  the  acid-neutralizing-  power  of  the  blood  ("alkalinity") 
is  increased  in  cancer;  this  is  probably  related  to  if  not  the  cause  of 
the  decreased  HCl  content  of  the  gastric  juice,  which  occurs  whether 
the  cancer  is  in  the  stomach  or  not.  As  this  alkalinity  is  not  associated 
with  an  increase  in  the  inorganic  bases  of  the  blood,  it  may  be  that  the 
proteins  have  an  increased  basicity.  However,  numerous  other  ob- 
servers describe  a  decreased  alkalinity  as  in  other  cachetic  conditions.®" 
The  blood  in  cancer  contains  less  calcium  than  normal,  which  results 
in  a  tendency  to  osteoporosis  ^  and  to  deposition  of  calcium  in  the  kid- 
ney ei)itlielium ;  ^'^  there  is  an  increase  in  the  i)otassium  of  both  the 
blood  and  tissues. - 

(7)  Immunity  Reactions  in  Cancer. — The  fact  that  a  certain  degree 
of  specific  innnunity  can  be  developed  against  normal  tissue  cells  (see 
Cytotoxins,  Chap,  ix),  has  encouraged  study  of  the  possibility  of  se- 
<;uring  immune  antibodies  which  might  be  specific  for  cancer,  and  has 
led  to  much  research  on  this  subject,^  with  results  as  yet  of  more  sci- 
entific than  practical  value.  There  is  no  doubt  that  the  body  has 
distinct  powers  to  inhibit  to  a  greater  or  less  degree  the  growth  of  tu- 
mors, and  to  destroy  many  of  the  cells  which  escape  from  cancers 
into  the  lymph  and  blood,*  while  in  experimental  animals  inoculated 
tumors  are  in  most  instances  unable  to  grow,  and  they  may,  after 
growth  has  'once  begun,  recede  or  even  disappear.  Furthermore,  ani- 
mals may  be  made  immune  to  tumors  to  which  they  would  otherwise 
be  susceptible.  Many  schemes  of  immunization  of  patients  by  injec- 
tion of  extracts  or  autolysates  made  from  their  own  tumors,  or  similar 
tumors  of  others,  have  been  tried  ;  but  in  the  hands  of  competent  and 
critical  observers  the  results  seem  to  have  been  practically  negative.'"' 
There  is  no  lack  of  evidence  that  cancers  do  produce,  in  greater  or  less 
amounts,  various  antibodies  of  some  degree  of  specificity  for  cancer, 
which  must  be  inteii^reted  as  evidence  that  cancer  proteins  are  in 
some  respects  different  from  the  normal  proteins  of  the  host ;  however, 
the  amount  and  specificity  of  these  antibodies  seems  to  be  low,**^  and, 
in  many  observations,  they  have  failed  to  be  demonstrated.  Indeed, 
Coca  in  his  review  states  unqualifiedly,  "The  usual  biological  tests  of 
complement  deviation  and  specific  precipitation  fail  to  show  the  hypo- 
as  Biocliem.  Jour.,  1906  (1),  207:  Watson,  Jour.  Path,  and  Baot..  1000  (1,3), 
429:  Sturrock,  Brit.  Med.  Jour.,  1913  (2),  780.  Tlie  OH  content  ot  the  blood  is 
constantly  increased  in  cancer  (^Tenten,  Jour.  Cancer  Res.,  1917  (2),  170). 
99  See  Traube.  Int.  Zeit.  Phvsik.-Chem.  Biol.,  1014  (1),  380. 
iGoldzieher,  Verb.  Deut.  Path.  Ges.,  1012  (15),  283. 
laM.  B.  Schmidt,  Verb.  Deut.  Path.  Ges.,  1913  (16),  329. 
2Mottram,  x\rch.  :\Iiddlesex  Hosp.,  1910    (19),  40. 

3  Literature  by   Coca,   Zeit.  InimunitJit.,    1912    (13),   525;    Kraus   et   al..   Wien. 
klin.  Woch.,  1911    (24),  1003. 

4  Reviewed  by  Wells,  Jour.  Amer.  Med.  Assoc,  1009    (52),  1731. 

4a  See    Blumenthal,    Zeit.    Krebsforsch.,    1914     (14),    491:    Bauer,    Latzel    and 
Wessely,  Zeit.  klin.  Med.,  1915   (81),  420. 

4b  See  Morgenroth  and  Bieling,  Biochem.  Zeit.,  1915   (68),  85. 


508  THE    CHEMISTRY    OF    TUMORS 

thetical  antibodies,  tliongh  a  distinct  cytotoxic  influence  can  be  demon- 
strated in  the  plasma  of  animals  of  foreign  species  that  have  been 
actively  immunized  against  a  tumor."  His  own  experiments  failed 
to  demonstrate  specific  complement-fixation  antibodies  in  patients  in- 
jected with  extracts  of  their  own  tumors.  Lewin  ^  also  fails  to  find 
conclusive  evidence  of  the  demonstration  of  specific  antibodies  in 
cancer,  yet  accepts  the  immunity  which  is  produced  by  injections  of  a 
virulent  cancer  material  as  an  active  immunity  dependent  upon  cancer 
antibodies.  It  may,  however,  depend  on  a  stimulation  of  the  local 
cellular  reactions  that  inhibit  cancer  growth.^"  Pfeiffer  ^  claims  to 
find  specific  anaphylactic  antibodies  in  the  blood  of  cancer  patients, 
but  this  has  not  been  confirmed  by  several  other  observers.^'^ 

V.  Dungern  ^  has  claimed  to  secure  positive  complement  fixation 
reactions,  partially  specific  for  cancer  and  benign  tumors,  by  using 
alcoholic  extracts  of  the  tumors  or  acetone  extracts  of  human  ery- 
throcytes as  antigen,  but  he  interprets  these  reactions  as  not  due  to 
specific  antibodies,  but  to  abnormal  products  of  metabolism.  The 
complement  content  of  the  blood  is  said  to  be  slightly  increased  in 
cancer  ( Engel )  ,*  but  there  is  nothing  characteristic  about  this.  Ascoli 
and  Izar  **  have  applied  the  meiostagmin  test  (g,  v.)  and  state  that  this 
gives  very  positive  results  in  determining  the  existence  of  cancer,  their 
work  having  been  corroborated  by  many  but  not  by  all  of  those  who 
have  repeated  it.-'^  Burmeister  ^'^  could  obtain  no  reliable  results  with 
the  epiphanin  reaction. 

Freund  and  Kaminer  ^^  have  found  that  the  serum  of  cancer  pa- 
tients is  unable  to  dissolve  cancer  cells,  as  normal  serum  does,  and 
even  protects  them  against  the  lytic  power  of  normal  serum.  The 
lysis  is  ascribed  to  a  non-nitrogenous  fatty  acid,  while  the  protective 
agent  of  cancer  serum  is  said  to  be  a  "nucleo-globulin"  which  is  in- 
creased in  the  serum  in  cancer.  They  also  find  that  cancer  extracts 
give  a  specific  turbidity  or  precipitation  with  cancer  serum,  which 
is  attributed  to  a  carbohydrate  content  of  the  extract.  According 
to  Kraus  and  v.  Graff  ^-  the  serum  of  full  term,  pregnant  women, 
and   normal  umbilical   cord   serum,  behave  like   serum   from   cancer 

5  Folia  Serologica,  1911    (7),  1013:  literature. 

saTvzzor,  Jour.  Cancer.  Res.,  1916  (1),  12.5. 

cWien  klin.  Wodi.,  1909    (22),  989;   Zeit.  Imiminitiit.,  1910    (4),  45.5. 

f.aSoe  Weil,  .Jour.  Exp.  Med.,  Oct.,  1913. 

TMiincli.  ined.  Wocli.,  1912  (59),  65,  1093  and  2854;  also  Rosenberg,  Dent,  iiied. 
Woch.,  1912    (38),  1225. 

sDeut.  nied.  Woch.,  1910  (36),  986.  Not  corroborated  I)V  Ordwav  and  Kcllert, 
Jour.  :Med.   Research,   1913    (28),  287. 

oMiinch.  med.  Woch.,  1910    (57),  2129;   Biocheni.  Zeit.,  1910    (29),  13. 

oaSee  Rosenberg,  Deut.  med.  Wocli.,  1913  (39),  926;  Wissung,  Rerl.  klin.  Woch.. 
1915    (52),  998.' 

10  See  Burmeister,  Jour.  Inf.  Dis.,  1913  (12),  4.59;  Bruiriremann,  ^Slitt.  (irenz. 
Med.  u.  C'hir.,  1913   (25),  877. 

11  Biochem.  Zeit.,  1912  (46),  470;  Wicn.  klin.  Wo.Ii.,  1911  (24),  1759;  1913 
(26),  2108. 

i^Wien.   klin.   Wodi.,    1911    (24),   191. 


CHEMISTRY  or  ('i:irr\i\  si-rr/ric  ri  uons  509 

patients,  lu  ibUpport  of  Freuiul  and  Kaniiner's  observation  is  the 
experiment  of  Neuberg  ^^  who  found  that  cancer  cells  plus  noi'mal 
serum  underwent  digestion  more  rapidly  than  cancer  cells  plus  cancer 
serum,  as  measured  by  the  incoagulable  nitrogen.  A  critical  test  of 
many  reconunended  methods  of  serum  diagnosis  of  cancer  by  Hal- 
pern  ^^  gave  disappointing  results.  With  the  von  Uungern  technic  he 
obtained  80  per  cent,  of  positive  results,  with,  the  meiostagmin  reaction 
85  per  cent.,  but  with  the  Abdei'halden  method  but  30  per  cent.  The 
other  methods  he  finds  of  little  value.  The  testimony  concerning  the 
specificity  of  the  Abderhalden  reaction  in  cancer  is  so  conflicting  that 
it  seems  unprofitable  to  discuss  it,  the  results  varying  from  such  as 
those  cited  by  Ilalpern  above,  to  100  per  cent,  correct  reactions  de- 
scribed by  others.  INfuch  weight,  however,  must  be  given  to  the  en- 
tirely unsuccessful  attempts  to  establish  the  principle  of  this  reac- 
tion with  refined  chemical  methods  by  Van  Slyke.^*''  Coca  '■^^  obtained 
entirely  unsatisfactory  results  with  both  the  von  Dungern  comple- 
ment fixation  test  and  the  Freund-Kaminer  reaction. 

]Many  observations  have  been  made  on  the  antitryptic  activity  of 
the  blood  in  cancer  (see  Chap,  iii)  which  has  usually  shown  an  in- 
crease (in  all  but  about  10  per  cent,  of  the  cases)  ;  but  many  other 
conditions,  especially  cachexia,  may  cause  positive  reactions.  Cancer 
serum  is  said  to  have  a  heightened  power  to  activate  pancreatic 
lipase.^^ 

B.  CHEMISTRY  OF  CERTAIN  SPECIFIC  TUMORS 

In  the  literature  are  to  be  found  a  few  studies  of  chemical  features 
of  some  forms  of  tumors,  which  may  be  briefly  discussed  to  advantage. 

(1)    BENIGN  TUMORS 

(a)  Fibromas  and  Myomas. — The  few  specimens  studied  show 
but  a  small  amount  of  nucleoprotein,  as  might  be  expected  from  the 
small  amount  of  their  nuclear  material.  Because  of  the  tendency  of 
fibromas  to  undergo  retrogressive  changes,  the  amount  of  calcium  is 
likely  to  be  large.  No  studies  as  to  the  special  features  of  their  col- 
lagen, as  compared  with  normal  connective-tissue  collagen,  seem  to 
have  been  made.  Lubarsch  ^^  found  no  glycogen  (microscopically) 
in  any  of  66  fibromas  he  examined.  Wells  and  Long  ^"  found  that  in 
uterine  fibro-myomas  but  one  per  cent,  of  the  total  nitrogen  is  purine 
nitrogen,  distributed  as  guanine,  44  per  cent, ;  adenine,  31  per  cent. ; 

i3Biocliem.  Zeit.,  1910  (26),  344. 

i4Mitt.  Grenz.  Med.  Cliir.,  1013  (27),  370.  See  also  Mioni,  Tiimrri.  1014  (3), 
697. 

i4aArdi.  Int.  :\Ied..  1917   (10),  .56. 

i*t(  Jovir.  Cancer  Research,  1917    (2),  01. 

15  Shaw-Mackenzie,  Proc.  Rov.  Soc.  Med.    (Pharmacol),  1012    (5),  152. 

leVirchow's  .Arch.,  1906   (183),  1S8. 

IT  Zeit.  Krebsforsch.,  1913   (12),  598. 


510  Till-:    Clll^MlSTh')     OF    Tl  MORS 

liypoxanthine,  25  per  cent.  The  relatively  large  proportion  of  pre- 
formed hypoxantliine  corresponds  with  the  abundance  of  this  purine 
free  in  unstriated  muscle.  Fibromyomas  are  able  to  deamidize  their 
guanine  and  adenine  to  xanthine  and  hypoxantliine,  and  contain 
guanase  but  not  adenase.  Extracts  from  uterine  fibromyomas  show 
practically^  the  same  composition  as  extracts  of  normal  uterus. ^''^ 

A  uterine  fibroid  analyzed  by  Becbe  ^*  contained  14.56  per  cent,  of 
nitrogen,  0.981  per  cent,  of  sulphur,  0.139  per  cent,  of  phosphorus, 
0.013  per  cent,  of  iron,  0.12  per  cent,  of  calcium  oxide,  0.44  per  cent, 
of  potassium,  and  1.115  per  cent,  of  sodium.  The  proportions  of  ni- 
trogen and  sulphur  are  high  as  compared  with  most  tumors;  the  phos- 
pliorus,  iron,  and  potassium  are  low,  corresponding  to  the  small 
amount  of  nucleoprotein  and  the  slow  rate  of  growth.  If  degenera- 
tion is  marked,  the  amount  of  calcium  is  greatly  increased.  Kraw- 
kow  ^"  found  a  trace  of  chondroitin-sulphuric  acid  in  a  uterine  fibroid. 
Lubarsch  found  glycogen  occasionally  in  richly  cellular  uterine  leio- 
myomas, and  in  the  vicinity  of  degenerating  areas;  however,  76  out  of 
85  showed  no  glycogen.  Pfannenstiel  -°  analyzed  the  alkaline  fluid 
of  a  cystic  fibromyoma,  which  coagulated  spontaneously ;  it  contained 
sugar,  but  no  mucin  or  pseudomucin.  The  cysts  were  dilated  lymph- 
spaces,  and  the  fluid  corresponded  to  lymph  in  composition.  A  simi- 
lar result  was  obtained  by  Oerum,-^  who  found  in  the  fluid  serum- 
albumin,  serum-globulin,  and  0.358  per  cent,  of  fibrin;  the  total  pro- 
teins constituted  6.3056  per  cent.  Sollmann  —  found  in  the  "colloid" 
of  a  cystic  degenerated  fibromyoma  both  pseudomucin  and  paramucin 
(see  "Ovarian  Cysts"),  which  differed  somewhat  from  the  same  sub- 
stances found  in  ovarian  tumors.  From  a  myxoma  of  the  back  Os- 
wald --'■'  obtained  a  mucin  with  the  following  elementally  composition : 
C,  50.82 ;  H,  7.27 ;  N,  12.24 ;  S,  1.19 ;  P,  0.25  per  cent.  This  differs 
from  other  mammalian  mucins  in  the  presence  of  phosphorus,  but 
Oswald  does  not  consider  this  a  contamination.  It  also  contained  12 
per  cent,  of  carbohydrate,  apparently  glucosamine. 

The  common  occurrence  of  marked  cardiac  weakness  in  patients 
with  uterine  fibroids  has  led  to  the  suggestion  that  in  the  fibroids  some 
toxic  product  is  formed  which  acts  on  the  heart,  or  that  both  the  fibroid 
and  the  heart  defect  might  result  from  a  common  cause.  The  experi- 
mental evidence  concerning  the  relationship  is  not  convincing,  and 
there  is  much  ground  for  the  belief  that  the  heart  suffers  from  the 
anemia  common  in  these  eases.^^     There  is  said  to  be  a  hemolytic  poi- 

i7aWini\vaif<T.  Arcli.  f.  ClvnJik..  191.3    (100),  i530. 
isAmcr.  Jour.  Plivsiol.,  1!)04    (12),  107. 
i!>Arch.  cxp.  Path!  u.  Pharm.,  1898   (40),  195. 
20  Areh.  f.  Ovn.,  1890  (38),  4(58. 
2iMalv's  Jahrosbcr.,  1884   (14),  462. 

22  Amcr.  Oynccol.,  1903   (2),  232. 

22a  Zcit.   i)li.VKi(.!.  ("hcni.,    1914    (92).   Ml. 

23  Sw  Jasciiko,  Mitt.  Crenz.  Mod.  u,  Cliir.,  1912  (lo).  249;  IMcCliini,  Suifr., 
Gvn.,  Olist.,  1914   (18),  180. 


(JllEMlaTRy  OF  BESIUS   TLMOJi.S 


511 


son,  a  lipoid  according  to  ]\Iurray,-*  formed  in  the  degenerating 
fibroids  whicli  causes  local  liemolysis  and  "red  degeneration,"  and 
there  are  cases  of  acute  aseptic  degeneration  of  til)roinyonias  wiiieli 
seem  to  have  caused  systemic  intoxication. 

(h)  Chondromas,  like  normal  cartilage,  always  contain  much 
glycogen  (Luharsch).  ^Alorner -"'  found  cliondroitin-sulphuric  acid 
in  several  chondromas  that  he  examined,  althougli  Schiuiedeberg  had 
failed  to  do  so. 

(c)  Lipomas  have  been  studied  by  Schulz -^  and  by  Jaeckle.-^ 
The  former  found  in  a  retroperitoneal  lipoma  75.75  per  cent,  of  fat, 
2.25  per  cent,  of  connective  tissue,  and  22  per  cent,  of  water.  Of  the 
fat,  7.81  per  cent,  was  in  the  form  of  the  free  fatty  acids  and  92.7  per 
cent,  as  neutral  fats.  The  fatty  acids  of  the  fat  consisted  of  65.57 
per  cent,  oleic  acid ;  29.84  per  cent,  stearic  acid ;  4.59  per  cent,  pal- 
mitic acid.  Cholesterol  was  only  qualitatively  demonstrable.  In  the 
(.•onnective  tissue  was  found  chondroitin-sulphuric  acid.  Lubarsch 
found  glj'cogen  in  lipomas  only  when  they  were  degenerated. 
Jaeckle  observed  the  formation  of  calcium  soaps  in  a  calcifj-ing  li- 
poma, the  calcium  being  distributed  as  follows :  calcium  soaps,  29.5  per 
cent. ;  calcium  carbonate,  28.61  per  cent. ;  calcium  phosphate,  41.89 
per  cent.  The  fats  of  lipomas  he  found  practically^  identical  with 
those  of  the  subcutaneous  tissues,  except  sometimes  for  a  deficiency  in 
lecithin,  as  shown  by  the  following  figures : 


Composition  of 

Fats  in — 

Subcutane- 

Lipoma 

Lipoma 

Lipoma 

ous  tissue. 

I. 

II. 

III. 

Eefraction.  at  40° 

50.6 

50.1 

50.9 

50.5 

Saponitication  number    . 

197.3 

197.7 

197.7 

195.9 

Eeichert-Meisser  number 

0.25 

0.33 

0.35 

0.35 

lodin  number 

63.7 

59.0 

64.0 

64.1 

Olein 

74.1 

68.6 

74.4 

74.5 

Oleic  acid        .... 

70.9 

65.7 

71.2 

71.3 

Acid  number  .... 

0.39 

0.31 

0.48 

0.67 

Free  acid 

0.196 

0.1.55 

0.24 

0.34 

Palmitic  acid 

18.5 

24.9 

18.5 

Stearic  acid    .... 

6.2 

5.1 

5.9 

Lecithin 

0.084 

0.015 

Cholesterol       .... 

0.32 

0.34 

Lipomas  are  able  to  hydrolyze  fats  and  esters,  their  lipase  behaving 
in  all  respects  like  the  lipase  of  normal  areolar  tissue.-'*  Lipoma  fat 
is  hydrolyzed  by  lipase  as  readily  as  is  nonnal  human  fat.     No  rea- 

24  Jour.  Obs.  and  Gvn.,  1910  (17),  534. 

25  Zeit.  phvsiol.  Chem..  1895    (20),  357. 
sePfluger's  Arch.,   1S93    (55),  231. 

2T  Zeit.  phvsiol.  Chem.,  1902   (36),  53. 
28  Wells,  Arch.  Int.  Med.,  1912   (10),  297. 


512  TllL    {JllEiUSTRY    OF    TLMORH 

son  for  the  reputed  unavailability  of  lipoma  fat  for  the  metabolism 
of  the  host  could  be  fouud.--' 

{d)  Ovarian  cyst  contents  have  been  studied  more  than  almost 
any  other  tumor  products,  because  in  their  gelatinous  or  slimy  sub- 
stance are  contained  numerous  interesting  forms  of  proteins,  many 
of  which  are  combined  with  carbohydrates  and  related  to  the  true 
miucins.  These  substances  are  frequently  referred  to  under  the  names 
of  pseudomucin,  paralbumin,  metalbiimin,  and  ovarian  ''colloid,"  and 
belong  to  the  class  of  '^ mucoids."  ^^  In  view  of  the  fact  that  the  flu- 
ids in  the  Graafian  follicles  of  the  ovary  do  not  contain  these  particu- 
lar forms  of  protein,  their  presence  in  cj^sts  derived  from  adventitious 
structures  (Pfliiger's  epithelial  tubes)  suggests  a  specific  form  of 
metabolism  on  the  part  of  the  epithelium  of  these  structures. 

Serous  cysts,  formed  by  dilation  of  Graafian  follicles,  usually  are 
small  in  size,  and  the  contents  resemble  those  of  the  normal  follicles 
(Oerum),^^  consisting  of  a  serous  fluid  with  a  specific  gravity  usually 
from  1.005  to  1.014  (occasionally  1.020  or  more),  and  containing 
1.0-4.0  per  cent,  of  solids.  Occasionally  in  these  cysts  the  contents 
become  solidified  through  absorption  of  the  water,  and  a  gelatinous  or 
glue-like  "colloid"  content  results.  Mucoids  are  never  present  (Pfan- 
nenstiel)."- 

Proliferating  cystomas  contain  the  peculiar  characteristic  mucoid 
proteins  mentioned  above.  Usually  the  contents  are  fluid,  but  of  a 
peculiar  slimy,  stringy  character,  due  to  the  mucoid  substance,  and 
often  opalescent  or  slightly  turbid.  The  specific  gravity  is  generally 
high — 1.015-1.030.  The  reaction  is  usually  slightly  alkaline  to  lit- 
mus, and  neutral  or  slightly  acid  to  phenolphthalein.  If  hemorrhage 
has  occurred  into  them,  the  fluid  is  discolored,  and  may  contain  blood- 
pigments  in  ciystalline  and  amorphous  forms.  Small  cysts  often 
show  a  condensation  of  the  proteins  into  a  semisolid  "colloid"  ma- 
terial, but  sometimes  their  contents  resemble  those  of  a  serous  cyst. 
Often  masses  of  proteins  fall  out  of  solution,  forming  yellowish  floc- 
culi  or  large  deposits  half  filling  the  cysts.  As  with  all  stagnant  flu- 
ids of  this  type,  cholesterol  crystals  are  frequently  found.  The  char- 
acteristic proteins  are  members  of  the  class  of  pseudomucins,  which 
are  constantly  present  (Oerum). 

2»  III  xanthoma  tuhvrosum  multiplex,  wliicli  shows  local  deposits  composed 
largely  of  cholesterol  esters  and  contains  also  ]>igment  with  tlie  properties  of  a 
lipochrome,  the  jjresence  of  hvper-choiesterolemia  is  dis])uted.  (  lvosenl)looni. 
Arch.  Int.  Med.,  IDl.S  (12),  .Siif) ;  Scliniidt,  Dermatol.  Zeit..  1!)14  (21),  1:57). 
Edsall  fonnd  the  comijosition  of  tlie  fat  in  the  fatty  txmiors  of  adipoNis  dolorosa 
but  little  different  from  tliat  of  normal  fat.  ( (,)uo{ed  hv  Dercum  and  ?ilc('arthy, 
Amer.  Jour.  Med.  8ci.,  1!)02    (124),  994.) 

30  Concernin{^  mucoids  see  Mann's  "Chemistry  oi  the  Proteins,"  llHttl.  iiji.  541- 
551. 

31  See  Malv's  Jahresbericht.  1884    (14),  4.59. 

32  Arch.  f.  Viyna-k.,  1890  (.•?S),407   (literature). 


CHEMISTRY  OF  BEXKIX   TUMORS  513 

Chemistry  of  the  Mucoids  of  Ovarian  Cysts. — Pseudormuin  lias  tlie  following 
■elfiucntaiy  compositiuii :  C,  4!).75;  II,  (i.ilS;  X,  10.2S;  S,  1.25;  0,  31.74  per  cent. 
( Hammarsten) .  In  eoimnon  with  the  true  mucins  it  yields  a  sugar-like  reducing 
body,  which  has  been  investigated  by  numerous  chemists  (Miiller,  Panzer, 
Zangerle,  Ix^vthes,  Neubcrg,  and  ileymann^s).  Panzer  considers  that  this  re- 
ducing substance  is  in  the  form  of  a  sulphuric-acid  compound,  similar  to,  but  not 
identical  with,  chondroitin-sulphuric  acid.  Hammarsten,  however,  did  not  find 
this  substance  constantly  present.  Leathes  determined  for  the  carbohydrate  group 
the  composition  C,;lL:,NOi„,  named  it  ''paramucosin,"  and  considers  it  a  reduced 
<^houdrosin  (which  is  the  carbohydrate  group  of  chondroitin-sulphuric  acid). 
Neuberg  and  lleymann  established,  however,  that  tlie  reducing  body  must  come 
from  chitosainin  (CjHi-iNO.-,) ,  and  do  not  consider  paranuicosin  a  constant  con- 
stituent of  ovarian  mucoids.  The  amount  of  reducing  substance  varies  greatly 
in  the  mucoids  found  in  different  cysts;  in  some  the  mucoid  yields  but  about 
3  to  5  per  cent.,  in  others  as  much  as  30  or  35  per  cent.,  of  reducing  substance. 

Psendomucin  dissolves  readily  in  weak  alkalies,  and  differs  from  true  mucin 
in  that  it  is  not  precipitated  by  acetic  acid,  and  from  the  simple  proteins  in  that 
its  solutions  are  not  coagulated  by  boiling.  With  water  a  slimy,  stringy  semi- 
solution  is  formed,  resembling  in  appearance  the  material  found  in  ovarian  cysts. 
Leathes  distinguishes  two  forms  of  ovarian  mucoids:  (3ne,  paramucin,  occurs  as 
a  firm,  jellj'-like  substance,  which  is  converted  by  peptic  digestion  into  the  easily 
soluble  pseudomucin.  Ovarian  "coUoid"  probably  consists  of  a  thickened  pseudo- 
mucin.  often  mixed  with  other  proteins.  Pfannenstiel  32  considers  the  "colloid" 
material  as  representing  a  modified  pseudomucin,  strongly  alkaline  and  relatively 
insoluble,  which  he  calls  "pseudo-mucin  /3."  He  also  describes  a  very  soluble 
mucoid  found  only  in  certain  ovarian  cysts,  naming  it  "pseudo-mucin  7." 

The  reasons  why  these  variations  in  the  pseiidomucins  exist  is  not 
understood ;  they  cannot  be  explained  as  due  to  variations  in  the  cell 
type  in  the  cyst  wall,  althoug-h  pseudomucin  is  probably  the  result  of 
true  secretion.  The  smallest  cavities  of  ovarian  cystadenomas  con- 
tain nearly  pure  pseudomucin,  which  presents  a  clear,  glassy  struc- 
ture ;  the  larger  the  cysts  become,  and  the  more  turbid  and  thinner 
the  fluid  is,  the  more  simple  are  the  proteins  it  contains.  True  mucin 
is  never  present  in  ovarian  cysts.  Pseudomucin  occurs  only  in  the 
glandular  proliferating  cystomas  and  the  papillary  proliferating 
cystadenomas,  in  the  former  appearing  constantly  and  abundantly,  in 
the  latter  not  constantly  and  never  abundantly  (Pfannenstiel) .  Paral- 
humin  (Scherer)  is  a  mixture  of  pseudomucin  with  variable  amounts  of 
simple  proteins.  Metalhumin  (Scherer)  is  the  same  body  that  is  called 
pseudomucin  by  Hammarsten.  Paramucin  (Mitjukoff)  ^^  is  a  mucoid 
differing  from  mucin  and  pseudomucin  in  reducing  Fehling's  solution 
directly,  without  having  the  carbohydrate  group  first  split  off  by 
boiling  with  an  acid.  Hj^drolysis  of  paranuicin  by  Pregl  ^''  showed  an 
absence  of  glycocoll,  but  traces  of  diamino-acids,  and  the  presence  of 
leucine,  alanine,  proline,  aspartic  and  glutamic  acids,  tryptophane 
and  tyrosine. 

Substances  similar  to  pseudomucin  have  been  occasionally  found  in 

cancerous  ascitic  fluid  and  in  cystic  fibromyoraas   (Sollmann)  ;  and 

they  are  abundant  as  constituents  of  the  contents  of  the  peritoneum 

33Hofmeister's  Beitr.,  1902   (2),  201   (literature). 
35  Arch.  f.  Gynaek..   1805    (40),  278. 
seZeit.  physiol.  Chem.,  1908    (58),  229. 

33 


514  THE    CHEMISTRY    OF    TLMOIiS 

in  the  condition  known  as  "pseudomyxoma  peritoniei,"  ^''  when  the 
material  is  in  reality  the  product  of  cells  implanted  on  the  peritoneal 
surface  through  the  bursting-  of  an  ovarian  cyst  (or  a  cyst  of  the  verm- 
iform appendix  (Friinkel)  ).^*  The  phj-sically  similar  substance  found 
in  pathological  sj^novial  membranes  by  Hammarsten  differs  in  yield- 
ing no  reducing  substance.  Parovarian  cysts  arising  from  the  Wolff- 
ian bod}'  present  an  entirely  different  content,  which  is  a  clear,  wa- 
tery fluid,  with  specific  gravity-  usually  under  1.010;  the  solids  amount 
to  but  1  or  2  per  cent.,  and  consist  chiefly  of  salts  (the  ash  being  often 
over  80  per  cent.),  mostly  sulphates  and  chlorides.  They  are  usually 
(or  always)  free  from  pseudomucin,  mucin,  or  other  sugar-containing 
substances,  and  other  proteins  occur  only  in  small  amounts,  unless 
the  cyst  is  inflamed.  Apparently  mucoids  do  not  form  in  cysts  lined 
b}'  ciliated  epithelium  (Pfannenstiel). 

Santi  ^^  has  studied  the  physical  chemistry  of  ovarian  cysts,  and  finds 
the  freezing  point  very  near  that  of  blood,  having  no  relation  to  den- 
sity, viscosity  or  nitrogen  content;  the  specific  electrical  conductivity 
is  higher  than  that  of  blood  serum.  The  physicochemical  properties 
are  less  dependent  upon  chlorides,  and  more  on  other  substances 
(Gnmer)." 

Intraligamentary  papillary  cysts  contain  a  yellow,  yellowish-green, 
or  brownish-green  liquid,  which  contains  little  or  no  pseudomucin ; 
the  specific  gravity  is  usually  high  (1.032-1.036)  and  the  fluid  con- 
tains 9  to  10  per  cent,  of  solids.  The  principal  constituents  are  the 
simple  proteins  of  blood  serum  (Hammarsten). 

According  to  the  same  author,  the  rare  iuho-ovarian  cysts  contain  a 
watery  serous  fluid  with  no  pseudomucin. 

(e)  Dermoid  cysts  of  the  ovary  contain,  as  their  chief  and  most 
characteristic  constituent,  a  yellow  fat,  which  melts  at  3-4°-39°  and 
solidifies  at  20°-25°.  Ludwig  and  Zeynek  *^  have  examined  over  sixty 
such  tumors,  and  found  that  the  fatty  material  constantly  contains 
two  chief  constituents :  one,  crystallizing  out  readily,  they  believed  to 
be  cctyl  alcohol, 

(CH3—  (CIL),,  — CILOII)  ; 

the  other,  remaining  as  an  oily  fluid,  seems  to  be  closely  related  to 
cholesterol,  although  not  consisting  of  one  substance  alone.  Small 
quantities  of  arachidic  acid  (C^nH^oOo),  as  well  as  stearic,  palmitic, 
and  myristie  acid  (Ci4H.,sOo),  existing  as  glycerides,  are  also  pres- 
ent. Ameseder,"*^  however,  found  evidence  that  the  supposed  cetyl  al- 
cohol   is    reall.y    eikosyl    alcohol     (CooH^oO).     Tliese    substances    are 

37  Litoraturo  l)v   Potcrs.   :\r()iiatsclir.   f.   Cch.   u.   C,\u..    ISOO    (10),   74!)-.    Wobor, 
St.   Pctorsb.   inod.'  Woeh.,   1001     (26),   XU. 
ssMiincli.  mod.  Woc-li.,  1901    (4S),  OfJf). 
3!'l''()]iii  dill,  cliiniica  ot  inicrosco]).,   1010    (2),   73. 

40  7{i()cli('in.   Jour..    1007    (2).   3S;{. 

41  Zoit.  jilivsiol.  f'licni..  1S07    (2.3),  40. 
*^  Jhid.,  1007    (.'")2),   121. 


CHEMISTRY  OF  MALIONAyT  TUMOh'S  515 

secreted  by  tlie  glands  of  the  cutaneous  structures  of  the  cyst,  and 
resemble  in  composition  sebaceous  material,  which  is  characterized  by 
containing  a  large  proportion  of  cholesterol  partly  combined  with  fatty 
acids.  Dermoids  sometimes  contain  masses  of  fatty  concretions  which 
seem  not  to  depend  on  chemical  changes  but  on  the  presence  of  forma- 
tive nuclei  and  framework  of  desquamated  epithelium ;  they  consist  of 
a  mixture  of  neutral  fats  and  cholesterol  esters,  with  some  free  cho- 
lesterol."-'^ Cholesteatomas,  in  addition  to  their  abundant  cholesterol 
content,  contain  keratin.'^ 

(/)  "Butter"  Cysts. — In  the  mammary  gland  retention  cysts 
form  filled  with  products  of  alteration  of  the  milk,  including  butyric 
acid 'and  lactose  (Klotz),"'"  and  these  are  called  "butter  cysts"  or 
milk  cysts.  Analysis  of  the  contents  of  such  a  cyst  by  Smita  *■'  gave 
the  following  results,  as  compared  with  human  milk : 


Fat  .     . 

Casein     . 
Albvunin 
Milk-sugar 
Ash 
Watei-     . 


Cyst  contents.  Human  milk 

72  07  3.nO 

.       4.37  0.63 

.        1.91  1.31 

0.88  6.04 

0.36  0.49 

.     20.81  87.09 


Fats  consisted  of —  , 

Cyst.  Cows'  milk. 

Stearin   and   palmitin 37.0  50.0 

Olein ^''•" 

Butyrin 9.0  7.8 

Occurring  independent  of  lactation  usually,  but  not  always,  are  the 
''soap  cystl"  which  contain  chiefly  calcium  and  magnesium  soaps, 
but   also   neutral   fats,   free   fatty    acids,    and   traces   of   cholesterol 

(Freund*''). 

(2)   MALIGNANT  TUMORS 

The  chief  general  features  of  the  composition  of  these  growths 
have  been  considered  in  the  discussion  of  the  chemistry  of  tumors 
in  general  (pases  494-509).  A  malignant  tumor  differs  from  a 
similar  benign  tumor  chiefly  in  having  usually  a  larger  proportion  of 
the  primary  cell  constituents,  and  a  smaller  proportion  of  the  sec- 
ondary constituents  and  intercellular  substances,  since  these  are 
largelv  the  product  of  the  functional  activity  of  the  cells,  which, 
in  malignant  tumors,  do  not  often  develop  sufficiently  to  functionate 
extensively.  Hence  malignant  tumors  usually  show  a  rather  high 
proportion  of  the  characteristic  constituents  of  nucleoproteins ;  i.  e., 
phosphonis  and  iron.     If  rapidly  growing,  they  contain  much  potas- 

42aLippert,  Frankf.  Zeit.  Path.,  ini3   (14),  477. 
43Risel.  Verh.  Deut.  Path.  Gesell.,  1900    (13),  322. 

44Arch.  klin.  Chir..  1880    (25),  40.  v,     •   i     ri     >, 

45Wien.   klin.   Woch.,    1800    (3),    551;    see   also    Zdarek.    Zeit.    physiol.    Chem., 
1008    (57),  461. 

46  Virchow's  Arch.,  1809  (156),  151. 


516  THE    CUEMIKTRV    OF    TUMORS 

sium;  if  iinderfjoin<y  much  retrogression,  little  potassium  and  a  larger 
amount  of  calcium  (Beebe,  Clowes  and  Frisbie).  On  account  of  the 
extensive  disintegration,  the  products  of  autolysis  are  usually  much 
more  abundant  than  in  benign  tumors.  The  composition  varies 
greatly  with  the  origin,  although  to  a  less  extent  than  with  the  benign 
tumors.  In  Fraenkel's  laboratoiy ''^  it  was  found  that  cancers  are 
often  defective  in  tryptophane,  and  from  a  squamous  cell  carci- 
noma of  the  skin  little  or  none  of  this  amino-acid  could  be  ob- 
tained, although  normal  squamous  epithelium  is  rich  in  trypto- 
phane. Fasal,*^"  however,  found  usually  a  high  tryptophane  figure  in 
cutaneous  epithelioma,  but  very  irregular  results  in  other  tumors. 
As  Bang  and  Beebe  have  shown,  the  tumors  arising  from  h'mphatic 
tissues  show  the  chemical  characteristics  of  these  structures,  and  con- 
tain histon  nucleinate.  Tumors  from  squamous  epithelium  develop 
keratin  in  direct  proportion  to  the  amount  of  maturity  the  cells 
reach.  Even  the  most  complex  and  specific  products  of  metabolic  ac- 
ti\aty  may  be  developed  by  malignant  tumors  (e.  g.,  thyroiodin, 
epinephrin,  bile),  and  in  a  form  and  condition  capable  of  performing 
function.  As  Buxton  has  shown,  malignant  tumors  produce  a  great 
variety  of  intracellular  enzymes.  The  idea  that  glycogen  is  present 
in  tumors  in  proportion  to  their  malignancy  has  been  disproved  by 
Lubarsch,  Gierke,  and  others;  among  the  malignant  tumors  glycogen 
is  found  particularly  in  chorioepitheliomas,  hypernephromas,  and 
squamous  cell  carcinomas.  Of  particular  importance  is  the  observa- 
tion of  Beebe,  that  the  composition  of  metastatic  growths  is  modified 
by  the  organ  in  which  they  are  growing,  so  that  they  tend  to  resemble 
the  organ  serving  as  their  host ;  which,  however,  does  not  hold  for 
certain  of  their  enzymes  (Wells  and  Long).  In  a  case  of  primary 
carcinoma  of  the  liver,  "Wolter  "^  found  the  tumor  tissue  richer  in 
nuelein  phosphorus  and  poorer  in  phosphatids  than  the  adjacent  liver 
tissue;  cholesterol  was  0.25  per  cent,  of  the  fresh  weight,  fatty  acids 
1.67  per  cent,  and  water  82.33  per  cent.,  the  water  of  the  normal 
tissue  being  79.34  per  cent. 

As  to  the  special  varieties  of  malignant  growths,  there  is  little  as 
yet  determined  concerning  their  chemistry  beyond  what  has  been 
stated  above.  Their  variations  in  composition  are  largely  the  direct 
result  either  of  their  resemblance  to  some  normal  tissue  or  of  degen- 
erative changes  that  they  have  undergone. 

"Colloid"  carcinoma  may  be  mentioned  specially,  in  view  of  the 
confusion  caused  by  the  lax  use  of  the  term  "colloid"  {q.  v.).  The 
fluid  contents  of  colloid  cancers  of  the  gastro-intestinal  tract  are 
usually  chiefly  epithelial  mucus,  containing  mucin  mixed  with  a 
greater  or  less  quantity  of  proteins  from  degenerated  cells  and  serous 

4TWion.  klin.  Woch..  1912   (25).  1041. 
47ar?iorhom.  Zoit,..  191.3    f.').5).  SS. 
ivb  Biochem.  Zoit.,  1913   (55),  260. 


{jUEMlHTUy  OF  MALltlXAM'  TUMORS 


517 


effusion.  Tliis  mucin  is  acid  in  reaction,  is  precipitated  by  acetic  acid, 
antl  has  an  affinity  for  basic  dyes/'"^^  The  colloid  cancers  of  the  mam- 
mary gland,  in  which  tlie  "colloid  degeneration"  involves  the  stroma, 
probably  contain  a  connective-tissue  mucin,  analogous  to  that  of  the 
umbilical  cord,  as  also  do  the  myxosarcomas,  if  we  may  judge  by 
their  origin  and  staining  reactions,  but  no  exact  chemical  study  of 
these  substances  can  be  found.  Colloid  cancers  of  the  ovary,  arising 
usually  from  the  same  structures  as  the  ovarian  cysts,  contain 
pseudomucin  or  allied  bodies  (see  "Ovarian  Cysts").  Colloid  tumors 
of  thj^roid  tissue  contain  the  typical  colloid  of  normal  thyroid  tissue, 
even  when  metastatic  in  other  organs;  in  the  tumor-colloid  is  a  rel- 
atively normal  proportion  of  iodin   (Gierke^®). 

Hypernephromas  possess  several  interesting  chemical  features. 
For  example  Gatti  ^°  brought  forward  the  fact  that  such  a  tumor 
analyzed  bj'  him  contained  3.4735  per  cent,  of  lecithin,  which  agreed 
very  well  with  the  amount  of  lecithin  in  normal  adrenals.  Beebe  ^^ 
found  in  the  watery  extract  of  a  hypernephroma  the  following  sub- 
stances: tryptophane,  proteoses,  glycogen,  leucine,  and  tyrosine,  in- 
dicating the  occurrence  of  autolysis.  About  29  per  cent,  of  fat  was 
present,  which  was  all  extractable  without  pepsin  digestion,  and  the 
fat  contained  about  18  per  cent,  of  its  weight  as  cholesterol.  Lecithin 
was  also  present,  but  not  quantitatively  determined.  A  study  of  the 
fats  and  lipoids  of  hypernephromas  and  other  tumors  gave  the  results 
shown  in  the  following  table :  ^^ 


Ether-soluble  material    

Cholesieiol,    %    total   dry   weight 

"  %   dry,  fat-free   substance 

"  %   ether-soluble  substance 

Lecithin,    %   total   dry   weight    

"  9c   dry,   fat-free   substance    .  . 

%  ether-soluble   substance..  . 


36.3 
7.6 
11.9 
20.6 
11.8 
18.4 
33.0 


Hypernephromas. 


1 

2 

3 

28.0 

33.0 

38.4 

4.6 

6.7 

8.7 

6.4 

10.0 

14.0 

16.9 

20.4 

22.9 

6.0 

9.0 

8.3 

8.3 

13.4 

13.4 

22.7 

27.5 

21.4 

85.0 
0.5 
3.3 
0.7 
2.0 

13.3 
2.4 


**-l  "3 

(._, 

o^ 

o 

^  -2 

03 

3A 

S-i 

o 

o  4 

CZZ 

C  i 

■3 -5 

■£« 

c3 

C3 

O 

O 

8.6 

21.4 

2.2 

0.9 

2.4 

1.2 

26.1 

4.3 

1.7 

0.7 

1.9 

0.9 

20.0 

3.0 

a  . 


14.5 
1.6 
1.9 

11.0 
6.2 
7.3 

39.8 


Hypernephroma  No.  1. — Typical  specimen,  with  the  usual  amount  of  hemorrhage  and 
necrosis ;  cells  much  vacuolated. 

Hypernephroma   No.   2. —  Similar  to  No.   1. 

Hypernephroma  No.  3. — Primary  growth  resembled  more  a  papilloma  than  an  ordinary 
hypernephroma  in  most  places;  no  vacuolization  of  cells,  little  necrosis,  and  no  heniorrhasro. 

Hypernephroma  No.  4. — Tumor  resembling  a  lipoma,  with  a  stroma  in  places  resembling 
a  fibrosarcoma  in  structure.  In  only  a  few  areas  were  cells  present  resembling  adrenal  tissue, 
most  of  the  tissue  resembling  fatty  areolar  tissue. 

47c  The  fluid  of  a  colloid  cancer  of  the  peritoneum  examined  hy  Hawk  contained 
a  protein  resembling  serosa  mucin,  containing  11.5  per  cent,  of  N  and  0.8  per 
cent,  of  S.   (McCrae  and  Coplin,  Amer.  Jour.  Med.  Sci.,  1916    (151),  475.) 

48  Hofmeister's  Beitr.,  in02   (3),  280. 

49Virchow"s  Arch.,    1S97    (150),   417. 

50  Amer.  Jour.  Physiol.,  1904    (11),   139. 

51  Wells,  Jour.  Med.  Res.,  1908  (12),  461;  see  also  Steinke,  Frankfurt.  Zeit. 
Path.,  1910   (5),  107. 


518  THE    CHEMISTRY    OF    TUMORS 

It  will  be  at  once  observod  that  tlie  two  typical  hypernephromas,  Xos.  1  and  2, 
show  a  marked  resemblance  to  the  normal  adrenal  in  the  proportion  of  fat  and 
lipoids.  (The  lower  figure  for  lecitliin  in  No.  1  probably  is  due  to  the  fact  that 
this  specimen  liad  been  preserved  longer  than  the  otliers.)  This  was  what  was 
to  be  expected  from  the  microscopic  resemblance  of  these  tumors  to  adrenal 
tissue,  and  corroborates  the  results  of  Gatti's  and  Beebe's  ol)serva.tions  on  iso- 
lated cases.  More  surprising  is  the  fact  that  e<iually  comparable  results  were 
obtained  in  the  hyporne|)liroma  (No.  .3),  which  contained  only  cells  free  from 
vacuolization  and  not  at  all  resembling  adrenal  cells.  From  this  it  may  be 
concluded  tliat  in  these  tumors  of  adrenal  origin  tiie  amoiuit  of  fats  and  lipoids 
present  cannot  be  estimated  from  the  degree  of  cytoplasmic  vacuolization  of  the 
cells,  or  the  extent  of  necrosis;  the  fatty  materials  are  an  intefiral  part  of  the 
cells,  present  in  them  as  an  essential  constituent  and  not  as  the  result  of  de- 
generation. 

The  results  of  analysis  of  two  carcinomas  and  a  sarcoma  indicate  tliat  the 
hypernephromas  are  peculiar  in  their  close  resemblance  to  adi'enal  tissue  in 
respect  to  fat  and  to  lipoid  c(mtent.  The  amoiuit  of  all  these  constituents  in 
these  three  tiunors  is  far  below  that  found  in  the  hypernepiiromas,  althougli  in 
the  carcinoma  of  the  breast  the  amount  of  simple  fats  is  relatively  large,  as 
might  be  expected  in  view  of  the  function  of  the  cells  from  which  it  arose.  It 
is  interesting  to  note  that  a  carcinoma  of  the  gall  bladder  shows  a  rather  high 
proportion  of  its  fatty  material  as  cholesterol,  for  this  observation  may  l)ear  a 
relation  to  the  well-known  tendency  of  the  epithelium  of  the  gall  bladder  to 
form  ciiolesterol.  The  large  proportion  of  lecithin  in  the  sarcoma  of  the  liver 
may  possibly  be  due  to  the  influence  of  tlie  soil  upon  which  the  tumor  was  grow- 
ing, but  we  need  more  information  concerning  the  lipoid  content  of  other  malig- 
nant tumors  arising  in  diflferent  sites. 

Renal  hypernephromas  reproducing  the  adrenal  cortex  in  struc- 
ture do  not  contain  epinephrine,^-  but  tumors  of  the  adrenal  arising 
in  the  medulla  may  do  so.^^  Microscopically,  hypernephromas  con- 
tain much  glycogen.  The  special  tests  for  hypernephroma  tissue 
recommended  by  Croftan  seem  not  to  be  specific.^* 

Melanotic  tumors  produce  melanin,  which  seems  not  to  differ  at 
all  from  the  melanin  found  in  normal  pigmented  structures.  Hel- 
man  ■'■'  found  as  high  as  7.3  per  cent,  by  weight  of  melanin  in  melano- 
sarcomas.  (See  also  Melanin,  p.  471,  and  Enzymes  in  Tumors,  p.  500. 
Concorning  Chloromas  ■'■""  see  p.  476. 

MULTIPLE  MYELOMAS  AND  MYELOPATHIC   "ALBUMOSURIA" 

Multiple  myelomas  are  of  particular  chemical  inten'st,  because  of 
the  appearance  in  the  urine  in  such  cases  of  the  peculiar  protein  first 
described  as  an  alhumose  by  Bence-Jones,^^  and  now,  because  of  lack 
of  grounds  for  its  definite  classification,  generally  known  as  the 
*'Bence-Jones  hody"  or  ''Bence-Jonea  protein."  Because  of  the  ex- 
tensive bone  destruction  there  is  also  an  excessive  excretion  of  cal- 

52  0reer  and  Wells,  Arch.  Int.  Med..  1000  (4).  2!)];  Brooks,  Jour.  Kxp.  :Med.. 
inil    (14),  .').')0:    Ciaccio,  Deut.  Zeit.  f.  Chir.,   1910    (104).  277. 

na  Wegelin.  Verb.  Deut.  Path.,  Ces..  1011    (15),  2r^r^. 

54  Koerber.  Virch.  Arch.,  lltOS    (102),  :ir,G. 

■"■.n  Arch,   internat.   Pharmacodyn.,    100:?    (12),   271. 

.'.."■.a  Metabolism  in  chloroma  does  not  ditVer  from  leukemia  (Sakaguchi.  Mitt. 
Med.  Fak.,  Tokio.   1014    (1.3),   lOS). 

50  For  literature,  see  Simon.  Ath.t.  .Tour.  ^led.  Sci..  l!t02  (123).  0.30;  Wclier 
et  al.,  ihid.,  1003  (126),  044;  Mollalf.  Lancet,  l'.i(),-|  (1),  207;  Hoscntiloom.  Bio- 
chem.   Bulletin,    1011    (1),   Kil. 


MYHI.OI'ATIIIC  MJll    \l(tsrh-/A  519 

eium,'^''  and  soinotiines  metastatic  calcilleation  may  occur.'""'  Tliis 
variety  of  tumor  differs  from  the  standard  types  of  malignant  tumors 
in  that  it  involves  the  marrow  of  many  l)ones  simultaneously,  in  a  very 
diffuse  nuuiner,  without  usually  giving  evidence  of  a  true  metastasis. 
Ill  many  respects  it  resembles  the  leukemias,  pseudoleukemia,  and 
chloroma.  and  it  is  extremely  uncertain  as  to  where  in  tlie  classifica- 
tion of  tumors  and  of  the  diseases  of  the  blood-forming  organs  this 
disease  should  be  placed.  Histologically,  the  tumors  show  evidence  of 
being  derived  from  the  specific  cells  of  the  marrow,  either  from  the 
plasma  cells  (Wright)  or  from  the  neutrophile  myelocytes  or  their 
predecessors  (IMuir).  Cases  of  myeloma  without  the  proteinuria 
have  been  described,  and  also  a  few  instances  of  the  presence  of 
apparently  tyjiical  Bence-Jones  protein  in  the  urine  without  myelomas, 
but  with  bone  carcinomas,  leukemia  or  chloroma.'"'*"^ 

Properties  of  the  "Bence=Jones  Protein." — Not  to  go  into  de- 
tails, which  are  given  in  the  literature  cited,  the  important  facts  con- 
cerning the  "'Bencc-Jones  protein,"  and  its  appearance  in  the  urine 
(''myelopathic  alhumosuria,"  Bradshaw),  are  as  follows: 

It  is  a  protein,  the  exact  nature  of  which  has  not  been  determined ; 
at  first  considered  an  albumose  because  of  its  peculiar  reactions  to 
heat,  its  nature  has  since  been  contested,  but  the  weight  of  evidence 
seems  to  be  in  favor  of  the  contention  of  Simon  that  it  is  most  closely 
related  to  the  water-soluble  globulin  of  the  blood.  In  certain  cases 
it  partly  precipitates  spontaneously  from  the  urine,^^  and  it  may 
crystallize  in  the  renal  tubules.^^''  Its  most  characteristic  properties 
are  the  following : 

The  coajyiilation  temperature  is  low,  varying  from  4n°-60°  in  various  cases, 
and  being  considerably  modified  by  the  amount  of  salts  and  urea  present  in  the 
solution.  Probably  the  protein  forms  a  molecular  compound  with  the  salts 
which  is  more  staV)le  at  100°  than  at  lower  temperatures   (Hopkins  and  Savory). 

In  many  cases  the  coagulum  is  redissolved  on  heating,  and  reappears  on  cool- 
ing, but  tins  characteristic  feature  is  not  always  present,  and  often  disappears  in 
cases  where  at  first  it  is  present. 

A  precipitate  is  formed  by  strong  (25  per  cent.)  nitric  acid,  which  disappears 
on  heating  and  reappears  on  coolina:.  Strong  hydrochloric  acid  causes  a  dense 
precipitate,  wiiieh  is  quite  typical    (Bradshaw). 

No  precipitate  is  produced  by  acetic  acid,  even  in  excess,  and  tlie  addition  of 
acetic  acid  to  a  hot  coagailated  specimen  causes  prompt  solution  of  tlie  coagulum. 

Unlike  albumoses,  this  substance  does  not  dialyze;  the  salt-free  solution  left 
in  the  dialyzing  bag  does  not  precipitate. 

A  purplish-violet  color  is  usually  given  with  the  biuret  reaction,  l)ut  it  may 
be  more  reddish  in  color,  especially  if  little  copper  is  present. 

•''•"■I  Blatherwick,  Amer.  Jour.  Med.  Sci.,  1010    (151).  432. 

50b  Tschistowitsch   and  Kolessnikofl",  Virchow's   Archiv.,   1000    (107),   112. 

sGc  Glynn  has  described  a  irlycoprotein  resembling  Maimer's  body,  in  tlie  urine 
during  myeloma  (Liverpool  ^letl.  C'hir.  .lour..  1014,  p.  82):'  A  crystallizahle  pro- 
tein, resembling  tlie  Bence-Jones  body,  has  been  found  in  the  urine  of  a  woman 
with  gastric  cancer  without  anv  bone  involvement  (Schumm  and  Kimmorle.  Zeit. 
physiol.  Chem.,  1014    (02),   1).' 

57  Rosenbloom,  Arch.  Int.  Med.,  1012   (0),  255. 

57aLoehlein,  Cent.  allg.  Path.,  1013    (24),  953. 


520  THE    CHEMISTRY    OF    TUMORS 

Sulpluir  is  readily  split  ofl'  liy  alkalies,  reacting  with  lead  acetate  to  produce 
lead  sulphide   (Boston). 

After  standing  in  alcohol,  by  which  llie  body  is  precipitated,  it  loses  its  solu- 
bility  (differing  in  this  respect  from  albuuiosc). 

As  to  the  exact  nature  of  the  body,  little  can  be  said  at  the  present 
time.  Since  protoproteoses,  deiiteroproteoses,  and  peptone  are  split 
off  on  digestion  with  pepsin,  the  molecule  is  evidently  larger  than 
that  of  any  of  the  albumoses.  The  Avell-purified  substance  is  free 
from  phosphoms,  and  hence  contains  no  nueleins ;  but  it  contains  con- 
siderable sulphur  (between  1  and  2  per  cent.),  which  is  readily  split 
off.  Like  casein,  it  contains  no  hetero-group  (lack  of  heteroproteoses 
on  digestion),  but  differs  in  containing  a  carbohydrate  group  (in  small 
amount)  and  in  the  absence  of  phosplionis.  On  hydrolysis  ^Nlagnus- 
Jjexy  ^s  obtained  glutaniinic  acid,  tyrosine,  and  leucine,  but  no  glyco- 
coll.  He  found  the  nitrogen  distributed  as  follows:  amid-nitrogen, 
9.9  per  cent. ;  humin-nitrogen,  9.8  per  cent. ;  diamino-nitrogen,  6.4 
per  cent. — which  last  was  composed  of:  histidine,  0.9  per  cent.; 
arginine,  2.4  per  cent. ;  lysine,  3.0  per  cent.  The  extensive  analytic 
studies  of  Hopkins  and  Savory  °°  show  that  the  amino-acid  grouping  is 
that  of  a  typical  protein,  with  a  high  proportion  of  aromatic  radi- 
cals, similar  proteins  not  being  found  in  the  tumors  or  muscles  of  a 
typical  case.  In  fact,  the  amino-acid  content,  as  given  below,  indi- 
cates that  Bence-Jones  protein  is  as  distinct  from  other  proteins  in 
chemical  composition  as  in  its  physico-chemical  properties.  The 
amino-acids,  in  round  numbers,  were  isolated  in  the  following  per- 
centage proportions  of  the  entire  protein :  Valine-leucine  fraction, 
14 ;  glutamic  acid,  8  ;  aspartic  acid,  2 ;  proline,  2.7  ;  phenylalanine,  4.8 ; 
tyrosine,  4.2 ;  tryptophane,  0.8 ;  cystine,  0.6  ;  arginine,  6 ;  histidine,  0.8 ; 
lysine,  3.7;  sulphur,  1.2.  An  important  point  in  this  work  is  the 
agreement  in  composition  of  the  proteins  from  two  different  cases, 
being  identical  within  the  limits  of  the  analytic  methods,  showing  that 
the  protein  is  of  constant  and  characteristic  properties. 

Occurrence  of  "Myelopathic  Albumosuria." — ^Not  all  cases  of  mul- 
tiple myeloma  show  the  presence  of  Bence-Jones  protein  in  the  urine, 
however,  and  it  is  present  occasionally  in  other  conditions.  Multiple 
bone  involvement  by  other  tumors  does  not  often  cause  "albumo- 
suria. ' '  *'°  There  is  no  evidence  that  it  occurs  in  the  normal  body,  even 
in  the  bone-marrow,  or  that  it  is  produced  as  a  step  in  the  splitting  of 
any  form  of  proteins.  A  few  cases  of  supposed  osteomalacia  have 
been  reported,  with  the  Bence-Jones  body  in  the  urine,  but  on  more 
careful  investigation  these  seem  to  have  been  unrecognized  myelomas 

58Zeit.  phvsiol.  Cheni.,  1000   '.30\  200. 

r-oJour.  of  Physiol.,  Iflll    (42).   180. 

60  A  case  of  this  kind  has,  however,  been  described  by  Oerum  (Ugeskrift  f. 
Lager.,  1004,  No.  24),  in  which  the  bone  tumors  were  multiple  metastases  of  a 
gastric  carcinoma.  See  also  Boggs  and  (inttiric,  .\mcr.  .Tour.  !\Icd.  Sci.,  1012 
(144),  803. 


MVKI.OI'ATIIir  ALIUWIOSI  in  A  521 

(e,  g.,  tlic  cases  of  Bence-Jones  and  of  Jochmann  and  Schumm). 
Similarly  the  case  reported  by  Askaiiazy  as  leukemia  with  Bence- 
Jones  protein  in  the  urine,  on  reexamination  was  found  to  be  nuiltiple 
myeloma.  However,  at  least  eight  cases  of  true  chronic  leukemia  with 
Bence-Jones  proteinuria  have  been  reported."*"'  Coriat  "^  describes 
a  substance  found  in  a  pleuritic  fluid  which  gave  the  reactions  of  the 
Bence-Jones  body,  and  he  believes  that  it  may  have  been  formed  from 
serum  globulin  through  the  digestive  action  of  the  leucocytes  or  bac- 
teria. Zuelzer  reports  finding  the  same  body  in  the  urine  of  a  dog 
poisoned  with  pyridin."-  It  is  a  striking  fact  that  the  kidneys  elim- 
inate such  great  quantities  of  this  protein  without  being  permeable 
to  the  very  similar  normal  blood  proteins,  and  usually  without  show- 
ing evidence  of  structural  changes.  It  may  be  found  in  the  blood  and 
exudates  of  patients  with  myeloma.*'-" 

Origin  of  the  Protein. — As  to  the  place  of  formation  of  this  pe- 
culiar protein,  there  is  much  diversity  of  opinion.  ]\Iagnus-Levy  ad- 
vanced against  the  idea  that  it  is  formed  b}'  the  tumor  cells,  the  fol- 
lowing arguments :  In  the  urine  of  myeloma  patients  are  excreted 
great  quantities  of  the  protein, — as  much  as  30  to  70  grams  per  day, 
■ — -whereas  the  total  amount  of  protein  in  all  the  tumor  tissue  in  the 
body  seldom  exceeds,  or,  indeed,  equals  this  quantity.  It  seems  im- 
probable that  so  little  tumor  tissue  can  form  so  much  urinary  protein, 
and  Magnus-Levy  suggests  that  it  must  come  from  the  food  proteins 
as  a  result  of  altered  protein  metabolism.  Against  this  view,  however, 
are  the  following  facts:  (1)  The  Bence-Jones  body  has  been  found 
(but  not  constantly)  in  the  myeloma  tissue,  but  not  in  other  organs 
or  tissues;  (2)  the  quantity  in  the  urine  is  not  dependent  upon  diet; 
( 3 )  it  is  associated  almost  exclusively  with  this  form  of  tumor.  Simon 
considers  it  probable  that  the  protein  is  formed  from  serum-globulin^ 
perhaps  by  an  enzymatic  action  of  the  tumor  cells,  and  once  formed, 
it  is  rapidly  eliminated  by  the  kidneys,  as  are  all  foreign  proteins. 
Normal  bone  marrow  does  not  contain  this  protein  (Nerking**^). 
Rosenbloom  '^^  has  found  evidence  that  Bence-Jones  protein  may  pos- 
sibly be  derived  from  the  osseo-albumoid  of  the  bones.  AYeber  and 
Ledingham  ^^  have  suggested  that  it  comes  from  the  cytoplasmic  resi- 
due of  karyolyzed  plasma  cells.  The  observation  that  under  benzol 
treatment  the  amount  of  Bence-Jones  protein  in  the  urine  of  leukemic 
patients  is  reduced  (Boggs  and  Guthrie  *"'-'')  is  also  good  evidence  of 
its  myelogenous  nature.     The  fact  that  Abderhalden  and  Rostoski  *"^ 

60a  Boggs  and  Guthrie,  Bull.  Johns  Hopkins  Hosp.,  1913    (24),  3G8. 
fii  Amer.  Jour.  Med.  Sci.,  1903   (126),  031. 

62Wolgemuth     (Arb.    a.    d.    Path.    Inst,    zu    Berlin.    Festschrift,    1906,    p.    627> 
states  that  normal  human  hone  marrow  mav  contain  true  albumoses. 
62a  Taylor  et  aJ.,  Jour.  Biol.  Chem.,  1917   (29),  425. 

63  Biochem.  Zeit.,   190S    (10),   167. 

64  Arch.  Int.  Med.,  1912   (9),  236. 

65  Folia  Hematol.,  1909    (8),  14. 

66  Zeit.  physiol.  Chem.,  1905   (46),  125. 


522  THE    CHL'MlsTh')     OF    TLMORS 

found  that  the  serum  of  rabbits  immunized  with  Bence-Jones  protein 
gives  the  precipitin  reaction  witli  human  serum,  is  evidence  that  the 
protein  is  a  human  tissue  protein  and  not  merely  an  absorbed  and 
excreted  food  protein.  This  has  been  corroborated  by  Hopkins  and 
Savorj-,*'^  who  also  found  that  the  amount  of  protein  in  the  urine, 
which  contained  about  one-third  the  total  nitrogen  excreted,  varied 
with  the  general  metabolism  and  was  not  controlled  by  the  diet. 
i\Iassini  "^^  reports  securing  positive  complement  fixation  tests  with 
immune  sera,  differentiating  the  Bence-Jones  protein  from  normal 
serum  proteins;  positive  sensitization  tests  were  not  obtained  by  cu- 
taneous injections  of  the  protein  by  Boggs  and  Guthrie.  Injected  into 
the  blood  it  is  non-toxic  and  does  not  lower  coagulability  as  a  proteose 
would.  It  is  capable  of  acting  as  an  antigen  in  anaphylaxis  reactions, 
wliich  also  indicates  that  it  is  a  complete  protein  and  not  a  cleavage 
product.®"  When  injected  into  dogs  it  is  partly  utilized,  although 
nephritic  animals  excrete  it  partly  hydrolyzed  into  proteose.*'-^ 

67  Corroborated  also  bv  Bosprs  and  Guthrie,  Amer.  Jour.  Med.  Sci.,  1012   (144), 
803;  Folin  and  Denis,  Jour.  Biol.  Chem..  1014   (18),  277. 
esDeut.  Arch.  klin.  Med.,   1911    (104).  20. 
69Tavlor  and  Miller,  Jour.  Biol.  Chem..  lOlG    (25).  281. 


C  PI  A  P  T  K  R    X  AM  1  I 

PATHOLOGICAL  CONDITIONS  DUE  TO,  OR  ASSOCI- 
ATED WITH,  ABNORMALITIES  IN  METABOLISM, 
INCLUDING  AUTOINTOXICATION 

During  the  course  of  metabolism  innumerable  organic  compounds 
are  formed,  some  of  which  are  of  a  more  or  less  poisonous  nature. 
As  long  as  the  body  is  in  a  nonual  condition,  these  injurious  sub- 
stances are  kept  from  accumulating  in  sufificient  quantities  to  do 
hann;  this  is  accomplished,  in  one  of  the  following  ways:  (1)  elimi- 
nation from  the  body  in  the  urine,  feces,  etc.;  (2)  combination  with 
other  substances  into  harmless,  or  relatively  harmless,  compounds;  (3) 
chemical  alteration  into  compounds  that  are  non-toxic  or  relatively 
innocuous.  Therefore  a  harmful  accumulation  of  metabolic  products 
may  be  the  result  of  any  one  of  the  following  conditions : 

(1)  Failure  of  elimination  because  of  abnormal  conditions  in  the 
eliminating  organs ;  e.  g.,  uremia. 

(2)  Failure  of  neutralization  by  chemical  combination,  presumably 
due  to  abnormalities  in  the  organs  or  tissues  through  whose  activities 
the  neutralization  is  normally  accomplished;  e.  g.,  diseases  of  the 
liver. 

(3)  Failure  in  the  chemical  transformation  of  the  metabolic  prod- 
ucts; this  may  result  either  from  abnormalities  in  the  functionating 
tissues,  or  through  a  checking  of  the  normal  steps  of  metabolism  by 
the  failure  of  elimination  of  the  end-products. 

(4)  Excessive  formation  of  certain  normal  products  of  metabolism; 
e.  g.,  hyperactivity  of  the  thyroid. 

(5)  Production  of  abnormal  toxic  chemical  substances;  e.  g.,  the 
intoxication  following  superficial  burns. 

Numerous  classifications  of  autointoxication  have  been  proposed  by 
various  authors,  some  excluding  from  the  causes  of  autointoxication 
all  but  the  products  of  metabolism  within  the  blood  and  tissues  of 
the  body,  as  has  been  done  in  the  preceding  consideration ;  many  in- 
cluding intoxications  caused  by  the  products  of  gastro-intestinal  fer- 
mentation and  putrefaction;  and  still  others  (v.  Jaksch)  including 
even  the  intoxications  produced  by  bacterial  invasion  of  the  body.^ 
It  is  extremely  difficult  to  draw  the  line  as  to  just  what  should  be 

1  See  resume  by  Weintraud,  Ergeb.  der  Path.,  1897    (4),   1. 

523 


524  ABWORMALITli:S    /X    METABOLISM 

included  under  the  temi  autointoxication,  and  particularly  difficult 
to  decide  the  proper  placing  of  the  intoxication  resulting  from  fecal 
retention  and  from  processes  of  decomposition  in  the  alimentary 
canal.  For  example,  the  poisoning  followinu,'  the  eating  of  partially 
decomposed  canned  food  could  not  be  looked  upon  as  an  autointoxi- 
cation, and  yet  there  is  no  fundamental  difference  whether  the  decom- 
position occurs,  as  in  this  case,  before  the  food  enters  the  body,  or 
whether  it  occurs  in  the  intestinal  tract  because  of  abnormal  bacteri- 
ological or  anatomical  conditions.  On  the  other  hand,  since  many 
of  the  obnoxious  products  of  metabolism  are  eliminated  through  the 
bowels,  failure  of  elimination  through  this  channel  may  lead  to  a  true 
autointoxication  as  much  as  may  deficient  renal  elimination.  On  the 
M'hole.  it  seems  best  to  restrict  the  term  autointoxication,  as  far  as 
possible,  to  the  disturbances  produced  by  products  of  metabolism 
that  have  been  formed  within  the  tissues  of  the  body  {intermediary 
metaholism),  considering  as  a  distinct  but  related  subject  gastro-in- 
testinal  autointoxication. 

In  the  discussion  of  autointoxication  from  the  standpoint  of  chem- 
ical pathology,  we  are  interested  particularly  in  the  chemical  nature 
of  the  substances  that  cause  the  intoxication,  and  in  the  chemical 
processes  by  which  their  action  is  kept  at  a  minimum,  rather  than 
in  the  clinical  features  or  anatomical  results  that  may  be  produced. 
Unfortunately,  in  but  a  few  instances  have  the  exact  chemical  sub- 
stances causing  these  intoxications  been  accurately  determined,  prob- 
ably because  in  most  cases  not  one  but  a  number  of  poisonous  sub- 
stances are  present;  and,  furthermore,  we  do  not  always  know  ex- 
actly when  a  certain  disease  is  to  be  ascribed  to  autointoxication,  nor 
can  we  always  determine  that  the  cause  of  a  certain  intoxication  lies 
in  an  abnormality  in  metabolism  and  not  in  an  infection  of  hidden 
nature.  It  is,  therefore,  quite  impossible,  with  the  uncertain  infor- 
mation available  at  this  time,  to  consider  autointoxication  in  a  sys- 
tematic way,  and  we  must  limit  ourselves  to  a  consideration  of  cer- 
tain pathological  conditions  in  which  there  appears  to  be  an  element 
of  abnormal  metabolism  with  resulting  intoxication.  In  some  cases 
this  intoxication  is  a  prominent  feature  of  the  disorder,  in  others  it 
is  subordinate  to  other  manifestations  of  the  disease ;  and,  finally,  we 
may  have  marked  alterations  in  metabolism  without  evidences  of  dis- 
turbance of  health   {e.  g.,  cystinuria,  alkaptonuria). 

Of  the  autointoxications  due  to  the  retention  of  poisonous  products 
of  metabolism  that  should  be  excreted  from  the  body,  first  in  order 
of  importance  stand  uremia  and  cholemia  (the  latter  has  already  been 
considered  in  connection  with  the  discussion  of  Icterus,  Chap.  xvi). 
Of  apparently  less  significance  are  autointoxications  due  to  failure  of 
elimination  of  gaseous  metabolic  products  by  tlie  lungs,  and  failure 
of  the  excretorv  functions  of  the  skin. 


UREMIA  525 


UREMIA  - 


The  cause  or  causes  of  the  severe,  often  fatal,  intoxication  that 
may  occur  when  the  outflow  of  urine  is  completely  checked,  or  when 
it  is  qualitatively  and  quantitatively  altered  for  long  periods  of  time, 
have  not  j^et  been  definitely  determined.  As  the  kidney  seems  to  be 
the  chief  organ  for  the  removal  of  the  products  of  nitrogenous  metab- 
olism, it  is  naturally  assumed  that  uremia  is  the  result  of  a  retention 
of  these  products,  but  as  yet  it  has  not  been  ascertained  which  of  the 
many  products  is  responsible,  and,  indeed,  there  are  very  good  rea- 
sons for  questioning  if  the  substances  present  in  normal  urine  do  or 
can  cause  uremia  when  their  elimination  by  the  kidney  is  defective. 
There  is  no  question  but  that  the  urine  contains  toxic  substances. 
Among  them  are  the  salts  of  potassium,  which,  however,  cannot  alone 
explain  all  the  urinary  toxicity,  for  the  symptoms  produced  by  the 
injection  of  urine  are  different  from  those  produced  by  potassium 
salts,  and  it  has  been  found  that  the  inorganic  constituents  (ash)  of 
urine  are  less  poisonous  than  the  entire  urine.  Furthermore,  toxic 
mixtures  of  organic,  ash-free  substances  have  been  obtained  from  nor- 
mal urine.^  Of  the  known  normal  constituents  of  the  urine  there 
-are  few,  however,  that  are  toxic  to  any  considerable  degree,  and  these 
•occur  in  but  very  small  quantities.  Urea  is  generally  considered  as 
almost  absolutely  non-toxic,  the  animal  body  withstanding  injection  of 
large  quantities  without  appreciable  injury.  Uric  acid,  the  purine 
bases,  hippuric  acid,  creatinine,  and  the  urinary'  pigments  are  all 
possessed  of  very  slight  toxicity,  and  their  effects  do  not  explain 
uremia.  Injections  of  urine  into  animals  may  cause  more  or  less 
disturbance,  but  it  is  different,  on  the  whole,  from  the  manifestations 
of  uremia.  (The  experiments  of  Bouchard  and  his  school  present 
such  serious  errors  of  technique  and  interpretation  that  they  are  now 
largely  disregarded.) 

For  these  and  other  reasons,  it  is  generally  considered  that  the 
intoxication  of  uremia  is  not  due  solely  or  chiefly  to  the  substances 
that  are  normally  eliminated  in  the  urine,  but  rather  to  more  toxic 
antecedents  of  the  nitrogenous  constituents  of  the  urine.  Urea  repre- 
sents but  the  final  product  of  a  long  series  of  reactions  by  which  the 
huge  protein  molecule  is  broken  up  into  its  "building-stones,"  the 
various  amino-acids,  and  these  in  turn  are  decomposed  in  such  a 
way  that  their  NHo  groups  are  combined  with  carbonic  acid  *  and 
eliminated  as  the  diamido-compound  of  carbonic  acid,  namely  urea, 

2  General  r^snin^  with  literature  bv:  Honigmann.  Ergeb.  der  Pathol.,  1894 
(Bd.  1,  Abt.  2),  639;  1902  (S),  .'549 •  Ascoli,  Vorlesimgen  iibcr  Uriimie.  Jena, 
1903. 

3  See  Dresbach.  Jour.  Exp.  Mod.,  1900    (."i).  31;"). 

4  Arginine  alone  of  all  the  amino-acids  splits  off  urea  directly  from  its  molecule. 


526  AB.\OiiUALlTlE>i    IS    METABOLISM 


0  =  c/         .     "We  know  that  the  liver  is  able  to  accomplish  the  con- 


,NH, 

\nil  • 

version  of  amino-aeids  to  urea,  for  it  has  been  experimentally  shown 
that  if  leucine  and  glycocoll  are  passed  through  the  vessels  of  the  iso- 
lated liver  they  disappear  in  part,  while  an  increased  amount  of 
urea  escapes  from  the  hepatic  veins.  It  is  probable  that  the  liver 
is  the  chief  site  of  urea  formation,  but  it  is  also  probable  that  urea 
can  be  formed  in  other  organs.  We  do  not  know,  however,  the  in- 
termediate steps  by  which  the  amiuo-acids  of  the  protein  molecule  are 
converted  into  urea.  It  has  been  repeatedly  shown  that  urea  can  be 
formed  from  ammonium  salts  of  organic  acids  (including  ammonium 
carbonate),  and  ammonia  is  a  constant  product  of  autolysis,  being 
characteristically  more  abundant  as  a  product  of  autolytic  proteolysis 
than  as  a  product  of  tryptic  proteolysis ;  therefore,  one  of  the  ante- 
cedents of  urea  is  probably  ammonia,  which  is  somewhat  toxic  and 
especially  hemolytic."  Another  antecedent  of  urea  is  ammonium  car- 
bamate, which  stands  in  structure  intermediate  between  urea  and  am- 
monium carbonate,  as  shown  by  the  following  graphic  formulae : 

,0H  .0  — NH,  .NH.  .NH. 

0=C<  0=C<  0=C<  0=C< 

(carbonic  acid)  (ammonium  carbonate)  (ammonium  carbamate)  .   (urea) 

That  ammonium  carbamate  is  possibly  an  important  precursor  of 
urea  has  been  shown  particularly  through  the  results  of  studies  of 
dogs  with  Eck's  fistula,*'  which  consists  of  a  fistula  between  the  portal 
vein  and  the  inferior  vena  cava,  the  blood  from  the  portal  system 
then  passing  directly  into  the  general  circulation  without  first  passing 
through  the  liver.  In  such  animals  the  urine  becomes  poor  in  urea 
and  relatively  rich  in  ammonium  carbamate.  At  the  same  time,  the 
dogs  show  severe  symptoms  of  intoxication  from  which  they  die,  and 
which  are  similar  to  the  symptoms  that  follow  intravenous  injection 
of  ammonium  earl)anmte.  Ammonium  carbamate,  being  a  substance 
of  considerable  toxicity  ^  when  free  in  tlie  bh)()d,  it  has,  therefore, 
been  quite  widely  considered  that  it  may  be  an  important  factor  in 
the  production  of  uremic  symptoms.  On  the  other  hand,  it  seems 
most  pr()ba])]e  that  the  condition  of  uremia  does  not  depend  upon  one 
but  upon  many  various  and  varying  su])stances,  especially  as  Hawk  '^ 
found  that  sodium  carbamate  did  not  produce  uremic  symptoms  in  his 
Eck  fistula  dogs,  while  Liebig's  extract  did."     Clinically  the  symptoms 

5  Concerning  the  toxicity  of  iunmoniuiii  sails  sco  IJacliford  and  Craiio.  'Medical 
News,   1902    (81),   778. 

6  See  Hahn,  Masson,  Xciicki,  and  rawlow,  Arcli.  f.  cxp.  l*alli,  u.  riiarm..  1S0.3 
(32),  161. 

7  See  Bickcl,  "Exp.  ITntersucli.  iibor  Cliolaciiiic,"  W'icsliadcii.  1000. 

8  Amor.  .Tour.  Pliysi(d.,   1008    (21),  2(10. 

0  Fisclilcr  l»(di('ves  tlie  iiiloxicatioii  wliicli  occurs  after  fcc(liii<r  meat  to  Eck 
fistula  dofis  to  be  an  alkalosis,  probably  from  Nil.,  salts  (Deut.  Arch.  klin.  Med., 
lit  11    (104),  .300). 


VRi:\iiA  527 

of  uremia  in  different  eases  are  widely  different;  thus,  if  uremia  is 
due  to  complete  suppression  of  urine  through  mechanical  obstruction, 
tlie  symptoms  are  quite  different  from  those  observed  in  the  uremia 
following-  a  chronic  nephritis;  drowsiness,  weakness  of  heart  action, 
and  syncope  being  the  chief  manifestations  of  obstructive  uremia, 
the  convulsions  and  other  manifestations  of  nervous  irritation  charac- 
teristic of  uremia  in  eliroiiic  nephritis  being  absent.'" 

Chemical  Changes  in  Uremia. — The  attempts  to  isolate  from  the 
blood  and  organs  of  uremic  patients  or  animals  toxic  substances  that 
explain  the  manifestations  of  uremia  have  thus  far  failed.  That 
there  is  an  actual  retention  of  organic  substances  in  the  blood  in  ure- 
mia is  shown  conclusively,  however,  by  the  studies  of  the  physico- 
chemical  properties  of  the  blood.  It  has  been  repeatedly  found  that 
in  uremia  the  freezing-point  of  the  blood  is  reduced  markedly  below 
the  normal; '^  instead  of  the  normal  depression  of  0.55°-0.57°  the 
freezing-point  is  usually  reduced  more  than  — 0.60°,  and  sometimes 
as  nnich  as  — 0.75°,  which  shows  that  the  number  of  molecules  in  the 
blood  is  increased.^-  At  the  same  time,  the  electrical  conductivity 
may  not  be  at  all  increased  (Bickel),'^  but  may  even  be  reduced; 
and  as  the  electrical  conductivity  of  the  blood  depends  upon  the  num- 
ber of  dissociable  molecules,  chiefly  inorganic  salts,  these  are  evi- 
dently not  increased.  Therefore,  the  increased  number  of  molecules 
must  represent  an  excess  of  organic  molecules  that  dissociate  but 
little  if  at  all,  and  hence  are  not  conductors  of  electricity.  Some 
authors,  indeed,  have  ascribed  uremia  to  the  increased  osmotic  pres- 
sure of  the  blood  from  the  retained  molecules,  but  this  is  improbable, 
according  to  Strauss,'*  who  found  that  a  marked  increase  in  molecu- 
lar concentration  may  occur  without  uremia,  and  that  we  may  have 
a  severe  uremia  without  increased  osmotic  pressure.'^ 

Careful  metabolic  studies  have  shown  that  nephritics  (chronic  inter- 
stitial) are  not  able  to  convert  proteins  into  urea  as  rapidly  or  as 
completely  as  normal  persons.'"  Erben  ''^  has  studied  the  variations 
in  the  normal  components  of  the  blood  during  nei^hritis.  and  found 
the  albumin  generally  decreased  in  proportion  to  the  globulin,  espe- 
cially in  cases  of  parenchymatous  nephritis;  lecithin  and  calcium 
are  also  decreased.     Rowe  '^'^  found  the  serum  proteins  greatly  low- 

10  Chiari,  however,  observed  true  uremia,  liotli  clinical  and  aiiatomioal.  in  a 
man  with  ureteral  obstruction   (Verh.  Deut.  Path.  Gosell.,  1!»12    [\'>) .  207). 

11  See  Tieken,  Amer.  Med.,  1905   (10).  pp.  .303.  .507,  and  822. 

12  See  table  of  freezinfj  points  of  blood  and  clfusions  on  page  35.5. 

13  Deut.  med.  Woch..   1002    (28),   501. 

1-4  Die  chronischen  Nierenentziindungen.  etc.,  Berlin,  1002. 

15  Stern  (Med.  Record,  1003  (63),  121)  notes  that  the  electrical  eondtictivity 
is  reduced  by  the  presence  of  excessive  quantities  of  non-electrolytes  in  uremia, 
and  regards  this  lowered  conductivity  as  a  factor  of  some  possiljle  importance. 

i«Levene  et  al..  Jour.  Exper.  :Med.,"  1000   (11),  825. 

I'Zeit.  klin.  Med..  1003    (50),  441:    1905    (57),  30. 

17a  Arch.  Int.  Med.,  1017   (19),  3.54. 


928  ABX0R1IAIJTIL\S    IX    METABOLISM 

ered  iu  chronic  nephritis  with  uremia,  an  increased  proportion  of 
globulin  being  present;  with  uremia  the  total  protein  content  is  nor- 
mal or  slightly  higher,  with  usually  increased  globulin,  while  nephritis 
without  edema  or  uremia  produces  a  marked  increase  in  the  globulin. 
The  decrease  in  red  corpuscles  and  hemoglobin  in  nephritis  is  a  well- 
known  feature. 

Orlowski  ^*  found  that  an  accumulation  of  acids  occurs  in  uremia, 
but  not  until  just  before  death,  and,  therefore,  the  reduction  of  blood 
alkalinity  is  not  the  cause,  but  an  accompaniment  of  the  uremia.  Fur- 
thermore, in  other  diseases  a  corresponding  or  greater  reduction  in 
alkalinity  may  occur  without  uremia.  Measurements  of  the  partial 
pressure  of  COo  in  the  alveolar  air  in  uremia  indicate,  however,  a 
certain  degree  of  acidosis.^''  This  seems  to  occur  to  a  sufficient  degree 
to  be  responsible  for  definite  clinical  symptoms  of  acidosis  only  in 
advanced  nephritis,  but  earlier  in  nephritis  an  acidosis  may  be 
demonstrable  by  the  alkali  tolerance  test  when  it  is  not  sufficient  to 
affect  the  alveolar  air.-°  The  development  of  this  terminal  acidity, 
together  with  the  finding  of  albumose  in  the  blood  of  a  nephritic  by 
Schumm,-^  suggests  the  probability  of  active  autolytic  processes 
occurring  in  uremia.  Neuberg  and  Strauss  --  have  also  found  glyco- 
coll  in  considerable  quantities  (1.5  per  mille)  in  the  blood-serum  of 
a  uremic  patient  and  in  the  blood  of  nephrectomized  rabbits.  The 
amount  of  colloidal  material  present  in  the  urine  is  decreased  in 
nephritis,  according  to  Pribram,-^  who  suggests  that  retention  of  this 
material,  which  is  rich  in  aromatic  radicals,  may  be  of  importance  in 
the  toxicity  of  uremia.  Rumpf  found  that  the  organs  of  nephritics 
contain  an  excess  of  potassium,  and  Blumenfeldt  -*  attributes  this  to 
a  defective  elimination  of  potassium  salts  which  he  observed  in  ne- 
phritis. 

Numerous  attempts  have  been  made  by  both  chemical  and  immu- 
nological methods  to  determine  wdiether  the  proteins  in  the  urine  in 
nephritis  come  from  the  food,  the  blood,  or  from  the  renal  cells  them- 
selves. In  alimentary  albuminuria  the  urinary  proteins  seem  not  to 
be  those  of  the  food,  but  human  proteins.-^  In  nephritis,  however, 
differentiation  between  serum  proteins  and  kidney  proteins  has  not 
yet  been  satisfactorily  accomplished.-" 

The  development  of  improved  methods  of  analysis  of  small  quan- 
tities of  blood,  and  other  fluids,  especially  by  Folin  and  Denis,  ]\Iar- 

isZontr.  f.  StofTwpchsol  \i.  Vcrdauunpskr.,  1902   (3),  123. 

19  Straub  and  Sclilavor,  Miinch.  med.  Woch.,  1912   (59),  5G9. 
20Peabody,  Arch.  Iiit.  INFod.,  1915   (16),  955. 

21  Hofmeister's  Bcitr.,   1903    (4),   4.53. 

22  13erl.  klin.  Woch.,  190G    (43),  258. 

23  Fortschr.  d.  Mod.,  1911    (29),  951. 
24Zeit.  exper.  PatlioL.  1913   (12),  523. 

25  Wells,  Jour.  Aincr.  ^Fcd.  Assoc.  1909    (.■)3),  803. 

20  Cameron  and  Wells,  Arch.  Int.  Med.,  lOl.'j   (l.")).  746. 


UREMIA  529 

shall,  and  \'aii  Slykc,  has  enabled  us  to  obtain  exact  knowledge  of 
many  of  the  chemical  changes  of  nephritis  and  uremia."^  It  has  been 
found  that  the  normal  blood  contains  from  20  to  30  mg.  of  nitrogen 
in  noncoagulable  form  in  each  100  c.c,  there  being  usually  about  5 
mg.  increase  after  meals,  and  ordinarily  about  one  half,  or  a  little 
more,  of  this  nitrogen  is  in  the  form  of  urea.  In  all  conditions  that 
impair  renal  function,  whether  renal  changes  or  circulatory  deficiency, 
there  is  a  rise  in  this  noncoagulable  nitrogen,  and  when  there  is  ex- 
cessive tissue  destruction  there  may  also  be  a  slight  rise  independent 
of  renal  injury.  As  a  general  rule,  but  with  some  exceptions,  the 
amount  increases  with  increased  renal  impairment,  the  highest  figures 
being  seen  in  uremia,  in  which  figures  as  high  as  350  mg.  have  been 
obtained.  Tn  130  uremics,  Foster  found  the  average  to  be  84  mg.  of 
nitrogen.  There  is  no  constant  relationship  between  the  blood  pres- 
sure and  the  nitrogen  figure,  but  functional  tests  usually  show  a  cor- 
respondence between  the  excretory  power  of  the  kidney  and  the  re- 
tention of  metabolites  in  the  blood.  The  symptoms  of  asthenic  uremia 
are  rarely  well  defined  when  the  concentration  of  urea  in  the  blood  is 
less  than  100  mg.  per  100  c.c,  and  thej^  are  rarely  absent  Avhen  the 
concentration  exceeds  200  mg.-* 

Along  with  the  other  nitrogenous  constituents  the  uric  acid  is  in- 
creased from  a  normal  2  to  3  mg.  up  to  7  to  10  mg.,  and  even  higher. 
Creatinine  rises  from  1  or  2  mg.  up  to  5  to  20  mg.-''  On  the  other 
hand  the  amino-acid  nitrogen  may  be  normal  in  the  blood  even  with 
extremely  high  nonprotein  nitrogen  figures,^"  although  sometimes  it  is 
much  increased,  as  high  as  30  mg.  amino  acid  N  having  been  found  by 
Bock  ^"^  in  uremia  (the  normal  figure  being  7  mg.).  Ammonia  nitro- 
gen may  show  a  slight  increase,  rising  in  half  of  Foster's  cases  from  the 
normal  0.5  mg.  to  from  0.7  mg.  to  2.2  mg.  per  100  c.c.  Indicanemia 
may  also  be  present,  but  it  is  not  a  toxic  factor.  (Dorner.)^^  The 
blood  normally  contains  about  0.05  mg.  per  100  c.c. ;  in  uremia  it 
may  rise  to  0.2  mg.,  and  as  much  as  2.2  mg.  has  been  found  in  one 
case.^- 

The  Etiology  of  Uremia. — The  fact  that  the  highest  figures  for 
non-protein  nitrogen  are  usually  found  in  uremia  might  be  accepted 
as  proving  that  uremia  is  caused  by  poisoning  with  these  metabolites, 
were  it  not  for  certain  contradictoiy  observations. 

27  Good  I'eviews  and  l)ihlioo;raphips  are  given  hv  Tileston  and  Comfort,  Arch. 
Int.  Med.,  1914  (14),  620;  Schwartz  and  :\rcGill',  ibid.,  1910  (17).  42:  Woods, 
ibid.,  1915   (16),  .577;  Karsner,  Jour.  Lab.  Clin.  :Med.,  1916   (1),  910. 

28  Hewlett,  Cxilbert  and  Wickett.  Arcli.  Int.  Med.,  191G   (18).  6.36. 

29  See  :Mvers  and  Fine,  Arch.  Int.  :Med  ,  1915    (16),  536;    1916    (17),  570. 

30  Foster,  Arch.  Int.  INled.,  1915    (15).  356. 
•"■oa  .Tour.  Biol.  Chem.,  1917    (29),  191. 

31  Deut.  Arch.  klin.  :Med.,  1914  (113),  342:  Rosenberg  Arch.  exp.  Path..  1916 
(79),  260:  TscherkotT,  Deut.  nied.  Woch.,  1914    (40),  1713. 

32Hass,  Deut.  Arch.  klin.  Med.,   1916    (119),   177. 
34 


530  ABNORMALITIES    IN    METABOLISM 

(1)  Occasionally  quite  typical  attacks  of  uremia  are  observed  with- 
out high  nonprotein  nitrogen  figures  for  the  blood,  even  as  low  as  28 
mg.  having  been  recorded  in  a  fatal  case.^^ 

(2)  Extremely  high  nonprotein  nitrogen  content  may  be  observed 
without  uremia.  Thus  Tileston  and  Comfort  found  169  and  150  mg. 
in  two  cases  of  acute  intestinal  obstruction  without  uremic  symptoms, 
and  similar  results  have  been  obtained  in  bicliloride  of  mercury  pois- 
oning,^* and  mechanical  anuria.  The  occurrence  of  albuminuric  re- 
tinitis also  seems  to  bear  no  relation  to  the  nitrogen  retention 
(Woods). 

(3)  None  of  the  known  nitrogenous  constituents  of  the  urine  can 
be  held  responsible  for  all  the  manifestations  of  typical  uremia  pois- 
oning. The  highest  purine,  uric  acid  and  creatinine  concentration  in 
a  given  case  may  occur  entirely  independent  of  uremic  couditions,^'^ 
the  amino-nitrogen  is  not  increased  in  uremia  and  urea  is  not  sup- 
posed to  be  toxic  in  this  degree.  To  be  sure,  an  unknown  toxic  sub- 
stance may  be  responsible,  but  in  some  cases  of  uremia  the  total  non- 
protein nitrogen  can  be  accounted  for  by  the  known  nitrogenous  com- 
ponents found  in  the  blood  (Foster). 

We  therefore  are  driven  to  one  of  the  following  alternatives : 

(1)  The  nerve  cells  may  be  made  hypersensitive  to  some  one  of  the 
known  constituents  by  the  excessive  amounts  of  the  other  metabolites. 
This  is  a  purely  speculative  hypothesis,  without  any  actual  evidence 
in  its  support. 

(2)  The  portion  of  unidentified  nitrogen  usually  present  in  the 
blood  may  contain  a  specific,  highly  efficient  poison. 

In  support  of  this  hypothesis  is  the  finding  in  a  series  of  cases  that  the  pro- 
portion of  noncoaguhible  blood  nitrogen  that  couhl  not  be  accounted  for  by  tlie 
known  nitrogenous  metabolites  seemed  to  vary  directly  with  the  severity  of  the 
symptoms    (Woods). 35a 

'  Ilartnian  3ob  has  suggested  that  the  substance  which  causes  the  characteristic 
odor  of  the  lu-ine  may  be  responsible  for  at  least  some  of  the  intoxication  of 
uremia.  This  substance,  which  he  has  isolated  and  described  under  the  name 
"urinod,"  he  believes  to  be  a  cyclic  ketone  with  the  empirical  formula  CgHsO; 
it  is  highly  toxic,  and  causes  mental  symptoms.  This  important  observation 
awaits  confirmation. 

Foster  s^r  has  described  the  finding  of  a  toxic  base  in  the  Ijlood  of  uremics, 
absent  from  the  blood  in  other  conditions,  whicli  causes  death  of  guinea  pigs  with 
symptoms  suggestive  of  tlie  eclamptic  type  of  uremia.  Further  development  of 
this  work  is  also  awaited. 

33  There  arc  few  who  would  go  to  tlie  extreme  of  Strauss  ( IJerl.  kliii.  Woch., 
1915  (.52),  .S08)  and  limit  tlie  term  lu-emia  to  cases  showing  a  liigli  non -protein 
nitrogen  in  the  blood,  no  matter  wliat  the  symptomatology  and  patliology  may 
be.  A  totally  difi"erent  view])()int  is  expressed  liy  iJeiss,  Zeit.  klin.  Med.,  1014 
(80),  97,  424,  4.')2. 

34  See  Foster,  Arch.  Int.  IVfed.,  l!)ir)   (],')),  754. 
35Mvers  and  Fine,  .Tour.  IJiol.  Cliein.,  1915   (20),  :V.n. 
35a  Arch.  Int.  Med.,   V.U',    (Hi),  .-)77, 

sab  Ibid.,  1915  (Ifi),  9H. 

30c  Trans.  Assoc.  Anier.    i'livs.,   1!)15    (.'JO),  305. 


UREMIA  531 

(3)   Uremia  may  not  depend  on  intoxication  of  tlie  nerve  cells,  but 
upon  the  nieclianical  effects  of  edema  involving  these  cells. 

One  of  the  striking  features  of  autopsies  of  uremics  is  often  the  ."wet  brain" 
and  the  excessive  amount  of  cerebrospinal  iluid  wiiicii,  during  life,  may  be  found 
lo  be  under  a  heightened  pressure.  We  l<nu\v  that  not  only  general  but  localized 
edemas  occur  in  nephritis,  and  tliat  localized  edema  in  the  brain  may  be  associ- 
ated Willi  and  apparently  responsible  for  paralyses,  convulsions,  hyperirritability 
and  mania.  The  wet  brain  of  nephritis  is  similar  to  the  wet  brain  of  acute 
alcoholism  and  delirium  tremens.  Oftentimes  the  nervous  symptoms  of  uremia 
are  distinctly  focal,  and  a  complete  hemiplegia  from  hemorrhage  may  be  exactly 
simulated;  convulsive  seizures  identical  with  those  of  brain  tumor  may  be  seen. 
It  is  extremely  dillicult  to  explain  these  localizations  by  the  action  of  a  soluble 
poison,  and  simple  if  we  assume  a  local  edema.  It  is,  of  course,  as  dilhcult  to 
explain  the  localization  of  the  edema,  but  we  know  that  in  nephritis  lucalized 
edemas  do  occur,  so  we  have  a  basis  for  the  assumption  of  localized  cerebral 
edemas.  A  general  acidosis  is  usual  in  nephritis  and  marked  in  uremia  as'i 
but  we  have  no  means  of  knowing  whether  local  acidosis  occurs  in  the  nervous 
system  that  may  be  responsible  for  local  edemas  according  to  Fischer's  hypothe- 
sis. Or,  osmotic  ell'ects  may  be  responsible,  in  view  of  the  demonstrated  high 
osmotic  pressure  of  the  blood  in  uremia,  and  the  fact  that  the  life  of  nephrecto- 
niized  rabbits  is  prolonged  by  giving  them  water.sse  In  any  event,  the  existing 
evidence  on  the  pathogenesis  of  uremia  does  not  explain  it  on  a  toxicologic  basis, 
and  hence  the  alternative  explanation  of  cerebral  edema  must  be  taken  into  con- 
sideration. 

On  the  other  hand  the  pathologist  recognizes  evidence  of  systemic 
intoxication  in  uremia.  The  uremic  pericarditis  and  endocarditis, 
which  have  often  failed  by  ordinary  methods  to  yield  bacteria,  are 
apparently  toxic  processes.  The  diphtheritic  colitis  indicates  vicari- 
ous excretion  of  poisonous  substances.  Structural  changes  are  found 
in  cells  that  suggest  poisoning ;  chromatolysis  of  the  cortical  ganglion 
cells  has  been  repeatedly  observed  in  uremia,  and  in  nephrectomized 
rabbits  Lewis  ^^^  found  acute  parenchymatous  and  fatty  degeneration 
of  the  myocardium  and  endothelial  cells  of  the  liver.  The  localized 
edemas  of  nephritis  often  show  a  fluid  of  the  character  of  an  exudate 
rather  than  a  transudate. 

It  would  seem,  despite  the  prevailing  opinion  to  the  contrary,  that 
it  is  entirely  possible  that  the  manifestations  of  uremia  may  be  caused 
by  the  known  nitrogenous  substances  that  the  kidneys  have  failed  to 
excrete,  and  that  the  only  difficult  thing  to  explain  is  the  failure  of 
investigators  to  consider  the  time  element  in  experimental  intoxica- 
tions. The  presence  of  200  mg.,  and  upwards,  of  nonprotein  nitrogen 
per  100  c.c.  of  blood,  which  is  often  found  in  uremia,  indicates  that 
the  blood  plasma  that  is  bathing  the  tissue  cells  contains  somewhere 
between  -0.5  and  0.7%  of  soluble  organic  substances,  a  strength  of 
solution  that  certainly  does  not  require  any  very  high  degree  of 
toxicity  when  continuously  maintained  at  this  concentration,  as  it  is  in 
nephritis.     The  reported  experimentally  determined   toxicities  with 

35d  Henderson,  Bull.  Johns  Hopkins  Hosp.,  1914  (25),  141;  Peabody,  Arch.  Int. 
Med.,  1915   (16),  955. 

35eCouvee.  Zeit.  klin.   Med..   1904    (54),  311. 
35f  Jour.  Med.  Res.,  1907   (17),  291. 


532  AliXORMALITlKS    1\    METABOLltiM 

these  substances  have  only  represented  transitory  conditions  which  are 
entirely  dissimilar  to  the  actual  conditions  in  the  body.  They  corre- 
spond to  the  cases  of  liigh  nonprotein  nitrogen  in  the  blood  in  in- 
testinal obstruction,  bichloride  poisoning,  etc.,  in  which  absence  of  tlie 
uremic  sj^mptom  complex  has  been  noted  and  remarked  upon.  To 
study  the  relation  of  uremia  to  retained  metabolites  we  need  observa- 
tions on  their  ettVcts  when  maintained  in  tlie  organism  for  long  periods 
at  the  concentrations  occurring  in  uremics  and  tliis  can  be  done  readily 
by  such  methods  as  have  been  devised  by  Woodyatt.^''^  A  start  in  this 
direction  is  furnished  by  Hewlett,  Gilbert  and  Wickett,-*  who  found 
that  when  large  doses  (100  to  125  gm.)  of  urea  were  given  to  normal 
men  there  occurred  symptoms  comparable  to  those  of  asthenic  uremia, 
wliich  appeared  only  when  the  urea  concentration  of  the  blood  had 
reached  levels  of  160  to  245  mg.  of  urea  per  100  c.c,  /.  e.,  just  the  con- 
centrations that  are  usually  seen  in  well  developed  uremia.  If  in  these 
experiments  of  brief  duration  such  marked  symptoms  were  produced 
by  urea,  what  striking  eifects  must  be  expected  when  these  same  urea 
concentrations  are  continued  in  the  blood  for  days  and  weeks  at  a 
time.  We  must  find  out  what  results  not  only  from  urea,  but  from 
creatinine  and  uric  acid  kept  in  the  blood  at  tlie  concentration  found 
in  uremia  for  long  periods,  as  well  as  any  other  substance  that  may  be 
increased  in  the  blood  in  uremia.  An  experiment  of  a  few  minutes' 
or  hours'  duration  cannot  be  expected  to  duplicate  or  elucidate  a  con- 
dition of  weeks  duration.  In  chronic  diseases  our  experimental  in- 
vestigations must  be  of  some  reasonably  comparable  duration,  and  this 
principle  of  investigation  is  now  made  possible  by  Woodyatt's  methods. 
And  finally,  in  view  of  the  extremely  varied  symptomatology  of  renal 
incompetence,  we  must  recognize  that  it  is  highly  probable  that  in 
different  cases  these  symptoms  vary  because  of  different  conditions. 
In  one  case,  urea  may  be  the  chief  factor,  in  another  the  action  of  urea 
Tnay  be  complicated  by  the  effects  of  acidosis  or  high  blood  pressure 
per  se,  while  in  others  cerebral  edema  may  be  the  chief  influence.  All 
possible  shades  of  cooperating  influences  may  be  expected  to  occur 
Avlien  the  kidneys  fail,  and  to  explain  the  confused,  variable,  changing 
picture  of  the  uremic  state.^^*^ 

35gJour.  Amer.  Mod.  Assoc,  191.5  (65),  2007;  Jour.  Biol.  Clieni..  1!)17  (20). 
355. 

35h  The  infliionoo  of  a  liypotlietioal  internal  socrotion  of  the  kidney  (Brad- 
ford), or  of  tlie  products  of  neplirolysis  (Asooli),  as  a  eause  of  uremia,  may  now 
ho  eonsidcred  as  of  historical  interest  only.  (See  Pearce,  Areli.  Int.  ^led.,  lOOS 
(2),  77;  1010  (.5),  133.)  Tlie  same  is  true  of  the  attempt  to  explain  the  liiirh 
lilood  pressure  as  the  result  of  adrenal  hvpertrophv.  (Pearce,  Jour.  Exp.  ^led., 
1908    (10),  735;   1910   (12),  128.) 


TOXKMIAS  or  rHEdXAXCY  533 


TOXEMIAS  OF  PREGNANCY  ^o 

Under  this  h('a(lin<«-  are  iiieliuled  eclampsia,  as  characterized  by 
convulsions  and  certain  anatomical  ohano-es,  together  with  those  in- 
stances of  intoxication  with  similar  anatomical  changes  and  no  con- 
vulsions, and  the  related  pernicious  vomiting  of  pregnancy.  Acute 
yellow  atrophy  of  the  liver  belongs  in  the  same  category,  although 
often  occurring  independent  of  pregnancy. 

ECLAMPSIA  " 

In  many  respects  eclampsia  resembles  uremia :  so  much  so,  indeed, 
that  Frerichs  and  others  have  referred  to  eclampsia  as  "puerperal 
uremia."  Considering  it  as  a  simple  uremia  occurring  in  pregnancy, 
uremia  and  eclampsia  have  in  common  the  constant  occurrence  of 
renal  disturbance  with  albuminuria  and  decreased  elimination  of  iTrea, 
and  also  violent  convulsions  and  profound  coma  teniiinating  in  death. 
On  the  other  hand,  eclampsia  differs  greatly  from  uremia  in  the 
anatomical  changes  observed  in  the  organs  of  the  body  other  than  the 
kidneys ;  these  are  of  such  a  nature  that  in  some  cases  it  becomes  diffi- 
cult to  distinguish  eclampsia  from  acute  yellow  atrophy  of  the  liver,^^ 
while  in  other  cases  the  picture  resembles  that  of  a  profound  bacterial 
intoxication,  so  that  numerous  authors  have  urged  that  eclampsia  is 
the  result  of  a  bacterial  infection.  At  the  present  time  the  cause  of 
puerperal  eclampsia  is  quite  unknown,  but  there  is  a  decided  ten- 
dency to  assume  that  poisonous  substances  are  developed  in  the  pla- 
centa or  fetus,  or  are  formed  in  the  body  as  a  reaction  of  the  maternal 
organism  to  the  foreign  fetal  elements.  These  theories  will  be  dis- 
cussed after  considering  the  known  facts  concerning  the  chemical 
changes  of  the  disease  that  have  been  reported  by  various  observers. 

Chemical  Changes  in  Eclampsia.— f/rinany  changes  are  practi- 
cally invariably  present,  and  usually  they  are  profound,  although 
there  are  no  known  characteristic  qualitative  or  quantitative  differ- 
ences from  the  urinary-  changes  of  puerperal  albuminuria  without 
eclampsia.  Proteins  are  abundant,  including  a  large  proportion  of 
globulin,  decreasing  rapidly  after  delivery  as  a  rule.  The  urea  is 
usually  very  low,  but  generally  increases  with  great  rapidity  after 
delivery,  until  two  or  three  times  the  normal  amount  is  passed  per 
day ;  as  urea  and  ammonia  do  not  seem  to  be  greatly  increased  in  the 
blood,  this  has  been  interpreted  as  indicating  that  during  eclampsia 

3«  Excellent  review  and  lnblioj:rapliy  by  Kwinjr,  Amer.  .Jour.  ^\ed.  Sei..  1010 
(130),  820. 

37  Literature  is  jriven  liv  Sikes  in  The  Practitioner.  100.5  (74),  pp.  47S  and 
642;  L.  Zuntz,  Handb.  d.  Biochem.,  1000,  HI  (I),  360;  Seitz,  Arch.  f.  Gyn.,  1000 
(87),  79. 

38  Concerning  the  liver  changes  see  Konstantinowitsch,  Ziegler's  Beitr..  1007 
(40).  483. 


534  ABXORMALITIES    IX    METABOLISM 

there  is  an  accumulation  of  tlie  precursors  of  urea  in  the  system 
(Sikes).  However,  the  involution  of  the  uteinis  itself  results  in  an 
increased  nitrogen  excretion  which  probably  accounts  for  much  if 
not  all  of  these  findings  (Slemons).^^^  There  is  an  excessive  elimina- 
tion of  nitrogen  in  the  form  of  ammonia,  which  is  said  to  be  due  to 
the  formation  of  abnormal  quantities  of  sarcolactic  and  other  organic 
acids  in  the  body,  which  are  combined  with  ammonia  in  the  blood  and 
eliminated  in  the  urine.^"  This  fact  has  led  many  to  look  with  favor 
upon  the  idea  that  eclampsia  is  due  to  an  acid  intoxication.  Other 
nitrogenous  urinary  constituents  may  also  be  increased,  so  that  the 
relative  proportion  of  nitrogen  eliminated  as  urea  is  often  greatly 
reduced.  It  is  said  that  the  toxicity  of  the  urine,  which  is  high  in 
normal  pregnancy,  is  increased  if  the  kidneys  are  not  impaired,  but 
decreased  if  their  permeability  is  impaired  by  nephritis,  the  character 
of  the  toxicity  being  such  as  to  indicate  that  it  is  from  substances  de- 
rived by  disintegration  of  proteins  (Franz).  The  proportion  of  sul- 
phur eliminated  in  an  unoxidized  form,  as  compared  with  that  elimi- 
nated as  SO4,  is  much  greater  than  normal.  These  findings  all  indi- 
cate that  oxidation  within  the  body  is  impaired.  There  is  more  or 
less  retention  of  chlorides,  but  there  is  nothing  characteristic  in  this.'*" 
In  spite  of  the  hepatic  lesions  of  eclampsia  the  tolerance  for  levulose 
was  not  found  impaired  by  Alsberg.*^ 

The  nonprotein  nitrogen  of  the  blood  is  but  little  increased  in 
eclampsia,  and  not  to  the  extent  usually  seen  in  uremia,  and  it  bears 
no  definite  relation  to  the  severity  of  the  symptoms  (Farr  and  Wil- 
liams)."^ They  found  from  25  to  72  mg.  per  100  c.c.  in  seven  cases. 
These  figures  can  be  reasonably  explained  as  the  result  of  tissue  dis- 
integration rather  than  renal  retention.  However,  Losee  and  Van 
Slyke  could  find  no  increase  of  amino-acids  or  other  intermediates 
of  protein  destruction  in  either  blood  or  urine  in  pregnancy  toxe- 
mias ;  ""  their  total  nonprotein  blood  nitrogen  figures  ranging  from 
25  to  46  mg. 

The  decrease  in  the  alkalinity  of  the  l)lo()d  observed  by  Zangmeister 
and  others  has  been  ascribed  to  the  formation  of  sarcolactic  acid  by 
Zweifel/-  who  failed,  however,  to  find  an  excess  of  CO.,,  or  to  detect 
oxybutyric  acid  or  oxalic  acid  in  the  blood.  As  to  the  blood  proteins, 
fibrin  has  been  found  increased  by  Kolman  and  by  Dieiist,'''  while 
Schmidt  found  a  relative  increase  in  tbo  gl()l)ulin.  Sikes  eoncludes 
that  the  statements  to  be  found  in  the  literature  eoncei'iiing  tlie  tox- 

38a  Bull.  Johns  TTopkina  ITosp.,  1914    (25),   iDfi. 

30  See   /wcifcl   and   Lockniann,  IMiincli.   med.    Worli.,    lOOli    dS).   ^HT ;    Cfut.   f. 
Gvn..   190!)    (:53),   847. 
■40  Zinsser,  Zeit.  f.  Gob.,  1912   (70),  200. 
41  Cent.  f.  C.yn.,  1910    (.34),  (>. 
4iaAmer.  .Tour.  j\Ied.   St-i.,   1914    (147),   or.fi 
••lb  Amor.  Jour.  ^Nled.  Sci..  1917    (15.3),  94. 
4-.iAroh.  f.  Gyn.,  1905    (7(5),  537. 
43  Arch.   f.  Gyn.,   1912    (96),   43. 


ECLAMPSIA  535 

icity  of  tlie  blood  in  eclampsia  leave  nothiiif^  proved  concerning  this 
point,  but  more  recent  studies  by  Graf  and  Landsteiner  *^  affirm  an 
increase  of  toxicity  of  the  blood,  not  due  to  any  special  poison  but  to 
an  increase  in  the  amount  of  the  toxic  substances  ordinarily  present. 
The  antitryptic  titer  of  the  blood  may  be  much  increased. ^^'^  Zang- 
meister  ^^  ascribes  importance  to  edema  of  the  brain,  liallerini  *" 
found  that  the  physico-chemical  changes  in  tlie  blood  are  quite  tlie 
same  as  in  corresponding  conditions  of  nephritis.  An  increase  in  the 
sugar  content  of  the  blood  has  been  observed  by  Benthin/^  but  no 
other  abnonnality  of  carboh^'drate  metabolism  is  usually  present. 
Blood  lipase  is  much  increased  because  of  the  hepatic  injury  (Whip- 
ple ).-''••' 

Theories  as  to  Etiology. — The  anatomical  changes  of  eclampsia 
are  such  as  to  leave  little  or  no  room  for  doubt  that  there  is  a  severe 
intoxication  with  poisons  that  have  a  markedly  toxic  effect  upon  all 
the  organs  of  the  body,  thus  differing  from  the  toxic  materials  at  work 
in  uremia,  which  seem  to  affect  chiefly  the  central  nervous  system. 
Repeated  bacteriological  and  histological  studies  have  failed  to  dem- 
onstrate that  infection  with  either  vegetable  or  animal  parasites  is 
the  cause,  and  clinical  observations  do  not  support  such  an  hypothe- 
sis. The  association  of  the  condition  with  pregnancy,  and  particularly 
the  rapid  improvement  that  often  follows  the  removal  of  the  con- 
tents of  the  uterus,  almost  compels  us  to  admit  that  the  causative 
agent  is  produced  by  the  fetus  or  the  placenta.  Some  investigators 
(Politi,  Liepmann)  believe  that  they  have  found  a  greater  degree  of 
toxicity  in  extracts  from  the  placentas  from  eclamptic  than  from  nor- 
mal women.  We  have  no  exact  ideas  as  to  the  nature  of  the  supposed 
toxic  substances,  except  that  recent  developments  in  the  study  of 
immunity  reactions  point  to  their  origin  from  proteolysis  of  tissue 
proteins,  presumably  from  the  placenta.  The  hypothesis  of  Zweifel 
that  lactic  acid  is  responsible  seems  untenable,  and  the  degree  of 
acidosis  present  is  not  sufficient  to  account  for  the  intoxication  (Losee 
and  Van  Sh-ke). 

The  Placenta  as  a  Source  of  Intoxication. — Histologists  having  fre- 
quently observed  placental  cells  in  the  blood  and  vessels  of  eclamptic 
patients,  it  was  once  suggested  that  multiple  capillary  emholi  of  pla- 
cental cells,  detached  from  chorionic  villi  and  forced  into  the  pla- 
cental circulation,  cause  the  manifestations  of  the  disease ;  this  theoiy 
is  entirely  inadequate,  however,  to  explain  all  the  features  of  eclamp- 
sia.    Eelated  to  this  hypothesis  is  the  idea  that  the  placental  tissues, 

"Cent.  f.  Gyn.,   1909    (3.3),  142. 
44a  Franz,   Arch.   f.  Gvn.,   1914    (102),  579. 
45Deut.  med.  Woch.,"l911    (37),  1879. 
40Annali  Ostet.  e  Gin.,  1910   (32),  273. 

4TMonats.  Geb.  u.  Gvn.,   1913    (37),  305;   Rvser,  Deut.  Arch.  klin.  Med.,   1916 
(118),  408. 
47a  Jour.  Med.  Res.,   1913    (24),  357. 


536  AIi.\URMALITIJ:,S    1\    inrrABOLLSM 

being-  foreign  to  the  maternal  organism  in  so  far  as  they  are  derived 
from  the  ovum,  give  rise  to  the  production  of  antibodies  {syncytioly- 
si)is)  by  the  mother,  which  are  toxic  for  pregnant  animals  (Ascoli), 
and  which  may  have  to  do  with  eclampsia  in  some  unknown  way. 
Kosenau  and  Anderson  found  that  guinea  pigs  could  be  made  anaphy- 
lactic to  guinea-pig  placenta,  showing  conclusively  that  the  placenta 
contains  i)roteins  foreign  to  the  motlier.  Attempts  to  establish  the 
anaphylactic  nature  of  eclampsia  have,  like  so  many  other  theories, 
foundered  on  the  fact  of  the  characteristic  anatomy  of  this  disease, 
wliich  is  never  seen  in  anaphylaxis.^'*  The  studies  of  Abderhalden 
have  shown  that  the  blood  of  ever}'  pregnant  female  animal  contains 
enzj'mes  which  have  a  specific  proteolytic  action,  and  so  the  possibility 
exists  that  abnormal  or  excessive  products  of  such  proteolj^sis,  or  a 
lack  of  adequate  defensive  digestive  action,  may  be  responsible  for 
the  toxemias  of  pregnanc3^  Esch  "  and  Franz  ^^  have,  indeed,  found 
evidence  of  the  presence  in  the  serum  and  urine  of  eclamptics,  of  sub- 
stances resembling  anaphylactic  poisons  in  their  action,  and  presum- 
ably derived  from  proteolysis  somewhere  in  the  body.  Franz  found 
That  if  the  poison  injures  the  kidneys  seriously  it  is  retained  in  the 
bod;,',  the  urine  ceasing  to  be  toxic,  wliich  has,  presumably,  a  relation 
to  the  toxicosis  of  eclampsia.^^ 

Liepman  "  and  others  have  reported  the  finding  of  a  considerable 
degree  of  toxicity  in  eclamptic  placentas,  but  this  is  probably  related 
to  the  increased  autolysis  observed  in  eclamptic  placentas  by  Dry- 
fuss.'*^  According  to  Mohr  and  Heimann,^''^  the  eclamptic  placenta 
shows  a  great  decrease  in  lecithin,  which  they  ascribe  to  the  increased 
autolysis,  and  to  the  hydrolyzed  lecithin  they  attribute  the  hemotoxic 
effects.  On  the  other  hand  JMurray  and  Bienenfeld  ''^  report  the  find- 
ing of  an  increased  amount  of  lipoids  in  eclamptic  placenta.''" 

The  Fetus  as  a  Source  of  Intoxication. — A  reasonable  view  of  the 
cause  of  eclampsia  is  that  it  is  initiated  by  the  excessive  products 

48  See  Felliinflcr.  Zoit.  Gelj.  u.  Gvn.,  1911  (68),  26;  :\rosbaclicr.  Dent.  med. 
Woch.,  1911  (37),  ]()-21.  However,  Vertes  (Monat.  (ieb.  u.  Gyn.,  1914  (40),  361, 
4(il' )  states  tliat  animals  dyiiij^  from  aiiajihylaxis  may  sliow  typieal  eelainptic 
tissue  clianges,  whieli  is  not  in  accordance  willi  tlie  observations  of  manv  otliers. 

49Miincli.  med.  Wocli.,  1912    (59),  461. 

50lhid.,  pajje  1702. 

51  Hull  and  Ehodenlmrfj  (Amer.  Jour.  Obst.,  1914  (70),  919)  ascribe  impor- 
tance to  leucine  derived  from  proteolysis  of  the  placental  elements,  while  Kiutsi 
(Zeit.  Gel),  u.  ^iyn.,  1912  (72),  57(i)  considei-s  the  nuclcins  of  the  {)lacenta  llie 
toxic  ajjents;    both   statements  beinfj  unconlirn\ed   and   improbable. 

■•sMiinch.  med.  Woch.,  1905  (52),  (;S7  and  24S4;  ]?oos,  P.oslon  .Med.  and  Surj;. 
Jour.,  1908   (158),  612. 

•"]?iochem.    Zeit.,    1908    (7),  493. 

5iJbid.,  1912    (4(i),  367. 

55Jour.  Obst.  and  Cvn.  Urit.  Empire,  1!»1()  (18).  225;  lii.nlicm.  Zcil..  1912 
(43),  245. 

•''«  The  hypothesis  of  ^lohr  and  I'rcund  llial  oleic  aejd  fidiii  llic  cehun])tic 
placenta  is  a  hcniohiic  fa<'1nr,  is  not  corroborated  bv  I'olano  (Zeit.  <ieb.  u.  Gvn., 
1910    (65),  581). 


JX'LAMI'SI.i  537 

of  riu'tabolisni  tlirown  into  the  blood  of  the  mother,  botli  from  the 
fetus  and  from  licr  own  overactive  tissues;  these  cause  injury  to  the 
kidneys,  k^adin":  to  a  further  retention,  or  injure  the  liver  so  that 
the  normal  metabolic  processes  of  that  organ  (particularlj'  oxidation) 
cannot  be  carried  on;  or,  perhaps  more  often,  both  liver  and  kidney 
as  Avell  as  other  organs  are  injured.  In  this  way  a  vicious  circle 
might  be  established  and  rapidly  lead  to  an  overwhelming  of  the  ma- 
ternal system  with  toxic  products  derived  from  both  her  own  and  the 
fetal  tissues.  It  must  be  admitted,  however,  that  the  rapid  improve- 
ment that  so  often  follows  removal  of  the  products  of  conception 
indicates  strongly  that  the  poisonous  substances  arise  chiefly,  if  not 
exclusively,  in  the  fetus  or  the  placenta.  But,  as  Liepmann  points 
out,  the  child  shows  relatively  little  evidence  of  intoxication,  while, 
on  the  other  hand,  eclampsia  may  develop  after  delivery  of  the  fetus, 
which  facts  speak  in  favor  of  the  place  of  the  origin  of  the  poison 
being  the  placenta  and  not  the  fetiLs,  and  death  of  the  fetus  seems  to 
have  no  effect  on  the  eclampsia.^'  Especially  important  in  this  con- 
nection is  the  observation  of  a  case  of  eclampsia  by  Hitschmann  ^^  in 
a  patient  with  a  hydatid  mole  and  no  fetus. ^'^ 

The  Ductless  Glands  in  Eclampsia. — In  view^  of  the  mystery  sur- 
rounding the  cause  and  effect  of  the  enlargement  of  the  thyroid 
during  pregnancy,  it  is  not  strange  that  the  suggestion  has  been 
made  that  the  enlargement  is  for  the  purpose  of  neutralizing  the 
excessive  amounts  of  toxic  materials  in  the  maternal  blood,  and  that 
failure  of  this  enlargement  is  responsible  for  eclampsia.  In  support 
of  this  idea  Lange  ^°  states  that  absence  of  the  normal  thyroid  en- 
largement is  usual  in  eclampsia,  and  Fruhinsholz  and  Jeandelize  ®^ 
note  the  frequency  of  eclampsia  in  myxedematous  women.  The  nota- 
ble influence  of  calcium  upon  convulsions,  and  the  possible  deficiency 
in  calcium  during  pregnancy,  has  led  to  the  suggestion  that  this 
may  be  responsible  for  eclampsia,®-  and,  since  the  parathyroids  are 
related  to  calcium  metabolism,  that  they  are  concerned ;  *'^  but  such 
theories  fail  to  explain  the  many  changes  other  than  the  con\-ulsions, 
and  have  not  been  accorded  much  importance.  Kastle  and  Healy  ^* 
consider  that  parturient  paresis  of  cattle,  which  bears  some  resem- 
blance to  human  eclampsia,  is  caused  by  absorption  of  toxic  substances 
produced  in  the  formation  of  the  colostrum;  it  is  cured  by  dilating 
the  lacteal  ducts  by  oxygen  or  other  means.     This  observation  lends 

5- See  Lichtenstein,  Zeit.  f.  G^ti.,  1912   (36).  1419. 
58  Cent.  f.  Gyn.,  1904   (28),  1*089. 

50  See  also  Gross  (Praper  mod.  Woch.,  1909  (.34),  36.5)  who  t'oinid  records  of 
seven  cases  of  eclampsia  with  hydatid  mole,  witli  or  without  a  fetus. 

60  Zeit.  f.  Geb.  vi.  Gvn..  1899  "(40),  34. 

61  Presse  Med.,  1902   (10),  1023. 

62  See  Silvestri,  Gaz.  desjli  Osped.,  1910  (31),  689;  Mitchell,  Med.  Record, 
1910  (78),  90G. 

63  Massacrlia  and  Sparapani,  Arcli.  ital.  Biol..  1907    (48),  109. 
6*  Jour.   Infec.   Dis.,   1912    (10),   226. 


538  ABXORMALITIE^    IX    METABOLISM 

support  to  the  tlieory  advaueed  by  Sellheim  ^'^  that  human  eclampsia 
is  of  mammary  o-land  origin. 

Pernicious  Vomiting  of  Pregnancy. — This  condition  is  inseparably 
associated  with  eclampsia  and  non-convulsive  toxemias  of  pregnancy, 
lliere  being  transitional  and  border-line  cases  of  all  sorts.  In  fatal 
cases  of  pernicious  vomiting  anatomical  changes  resembling  those  of 
eclampsia  have  been  found,  and  albuminuria  and  icterus  are  often 
observed.®^  The  chief  chemical  interest  in  these  cases  lies  in  the 
urinaiy  findings,  there  being  commonly  observed  a  relatively  high 
proportion  of  ammonia  and  undetermined  nitrogen  with  decreased 
urea,  which  findings  have  been  considered  indicative  of  defective 
oxidation  or  deaminization  (Ewing  and  Wolf)  and  of  prognostic  and 
diagnostic  significance  (Williams).  Underhill  and  Rand*'^  hold  that 
the  urinary  changes  are  entirely  compatible  wdth  those  which  can  be 
produced  by  starvation  which  is  present,  of  course,  in  pernicious 
vomiting ;  but  Ewing  ***  contends  that  there  are  other  underljdng 
factors  beyond  those  of  starvation. 

Summary. — ]\Iost  of  the  facts  at  hand  speak  against  the  idea  that 
one  definite  chemical  substance  is  responsible  for  the  anatomical 
changes  and  symptomatic  manifestations  of  eclampsia.  j\Iore  prob- 
ably there  are  present  not  only  the  poisonous  substances  that  initiate 
the  tissue  changes,  and  which  probably  originate  in  the  placenta 
itself  or  from  digestion  of  placenta  proteins  in  the  maternal  blood 
or  organs,  but  also  toxic  substances  that  accumulate  because  of  the 
disorganization  of  the  liver  and  kidnej^  cells,  and  which  are  possibly 
similar  to  the  toxic  substances  most  prominent  in  uremia  and  in  acute 
yellow  atrophy,  for  eclampsia  seems  to  stand  intermediate  between 
these  two  diseases,  encroaching  upon  the  _  characteristics  of  each. 
Acid  intoxication,  which  undoubtedly  exists  to  a  greater  or  less  de- 
gree in  some  cases  of  eclampsia,  is  not  an  important  cause  of  the  clin- 
ical manifestations  of  the  disease.  The  finding  of  minute  quantities 
of  lactic  acid  in  the  blood,  urine,  and  in  the  cerebrospinal  fluid  (Fiitli 
and  Lockemann)  is  not  of  great  signiticanee,  for,  as  Wolf""  rightly 
insists,  similar  amounts  may  be  found  in  other  conditions  associated 
W'ith  convulsions  and  partial  asphyxia,  or  in  partial  starvation,  such 
as  results  from  the  vomiting  of  pregnancy.  The  excretion  of  these 
organic  acids,  as  well  as  the  large  proportion  of  unoxidized  sulphur 
in  the  urine,  indicates  that  incomplete  oxidation  is  an  important 
feature  of  eclampsia,  and  under  such  conditions  a  large  number  of 
imperfectly  known  toxic  substances  may  accumulate  in  the  blood 
and  tissues.  The  defective  oxidation  and  deaminization  indicated  by 
the  urinary  findings  are  probably  the  result  of  the  injury  to   the 

osZent.  f.  Gyn.,   1909    (:U),   1G09. 

o'iSee  Kwinof' and  Wolf,  Anier.  Jour.  Ohslr.,   1907    (55).  2S9. 

87  Arch.  Int.  Med.,   1910    (5),  Gl. 

osAmr-r.  Jour.  Med.  Sci.,   1910    (130),  828. 

CO  New  York  Med.  Jour.,  190(i   (83).  813. 


ACUTE   YELLOW  ATROl'IIY  OF  THE  LI V Eli  539 

liver-cells,  which  have  such  a  i)n)iiiiueut  oxidizing  function.  The 
hypotheses  Avhich  ascribe  the  intoxication  to  products  of  specific  pro- 
teolj^sis  of  the  foreign  proteins  of  the  placenta  which  have  entered 
the  maternal  organism,  are  suggestive,  but  as  yet  are  not  sufficiently 
developed  to  permit  of  any  definite  conclusions  as  to  the  extent  to 
which  they  apply. 


ACUTE  YELLOW  ATROPHY  OF  THE  LIVER 

In  this  condition  there  is  presented  a  striking  picture  of  autolysis, 
in  that  a  large  parenchjTuatous  organ  undergoes  a  rapid  reduction 
of  size  because  of  a  solution  of  its  structural  elements,  while  at  the 
same  time  products  of  protein  digestion  (leucine,  tyrosine,  etc.) 
appear  free  in  the  liver,  the  blood,  and  the  urine.  Because  of  these 
prominent  features  and  their  relation  to  the  questions  of  metabolism 
in  general,  and  the  function  of  the  liver  in  particular,  acute  yellow 
atrophy  of  the  liver  has  been  the  object  of  much  greater  interest  and 
investigation  than  its  clinical  importance  would  warrant,  for  it  is  a 
rare  disease,  there  probably  being  but  about  500  cases  reported  in  the 
literature  to  1903,  according  to  Best's  figures."'' 

The  etiology  of  the  disease  is  quite  unknown,  but  it  is  very  prob- 
ably not  a  specific  one,  for  we  find  that  numerous  forms  of  intoxi- 
cation may  lead  to  a  condition  closely  resembling  acute  yellow  atro- 
phy,'^^  particularly  phosphorus  poisoning,  chloroform  poisoning, 
puerperal  eclampsia,  and  some  cases  of  septicemia  (especially  with 
the  streptococcus).'-  arsenic  poisoning  and  mushroom  poisoning."^ 
It  seems  probable  that  any  poison  which  does  not  directly  cause 
death,  but  which  causes  a  severe  injury  to  the  liver-cells  without  at 
the  same  time  destroying  the  autolytie  enzymes,  so  that  the  cells 
die  and  undergo  rapid  autolysis,  may  produce  a  condition  identical 
with  or  similar  to  acute  yellow  atrophy  (Wells  and  Bassoe).'"*  In 
the  typical  cases  of  the  disease,  of  "idiopathic"  origin,  the  poison- 
ous agent  possibly  comes  from  the  alimentary  canal,  as  indicated  by 
a  preliminary  period  of  gastro-intestinal  disturbance  that  usually  pre- 
cedes the  onset  of  the  disease,  and  secondly  by  the  fact  that  the  liver 
seems  to  receive  the  chief  effect  of  the  poison.  Whether  these  hypo- 
thetical poisons  are  produced  by  abnormal  fermentation  and  putre- 
faction in  the  alimentary  tract,  or  by  a  specific  organism  elaborating 
its  poison  in  this  location,  is  quite  unknown.  Bacteriological  studies 
of  the  disease  have  so  far  given  inconstant  and  non-instnictive  re- 

"0  Thesis,  University  of  Cliieaso,   1903. 

"1  It  is  to  be  borne  in  mind  tliat  tlie  color  is  yellow  only  during  the  earlier 
stages,  "red  atrophy"'  occurring  later,  but  the  name  acute  "yellow  atrophy"  has 
come  through  usage  to  ap]>ly  to  tlie  disease  as  a  whole. 

"2  Babes,  Ann.  Inst.  Path.  Bucarest,  vol.  6. 

'3  Frey,  Zeit.  klin.  Med..  1012    (75).  4.'}.5. 

'*  Jour.  Amer.  Med.  Assoc,  1904   (44),  685. 


540  AHSOiniAlATIES    1\    .UI:TA  HOLISM 

suits.  In  the  countries  wliere  phosphorus  poisoning-  is  common  (es- 
pecialh'  Austria)  there  has  been  found  much  difficulty  in  distin- 
guishing in  many  cases  the  results  of  phosphorus  poisoning  from 
acute  yellow  atrophy  of  the  liver,  and  many  have  contended  that 
there  is  no  real  difference ;  i.  e.,  that  phosphorus,  as  well  as  unknown 
poisons,  may  cause  acute  yellow  atrophy.  The  present  trend  of  opin- 
ion, however,  seems  to  favor  the  view  that  there  is  a  primary  liver 
atrophy  which  is  different  from  that  caused  by  phosphorus  or  other 
known  poisons  in  several  essential  respects.''* 

Phosphorus  Poisoning. — Between  phosphorus  poisoning  and  ^"pri- 
mary" hepatic  atrophjj  the  following  chief  differences  may  he  dis- 
cerned: Phosphonis  produces  a  general  injurious  effect  upon  all 
the  organs  of  the  body,  the  liver  merely  showing  the  most  marked 
anatomical  changes,  which  at  first  consist  of  a  fatty  metamorphosis 
of  the  liver,  due  to  migration  of  the  body  fat  from  the  fat  deposits 
into  the  injured  cell  (Rosenfeld,  Taylor)  ;  subsequently  the  liver 
cells  disintegrate,  the  cytoplasm  being  aft'ected  before  the  nucleus, 
and  the  liver  may  become  smaller  than  normal,  although  it  is  usu- 
ally enlarged  because  of  the  fat  deposition.  Typical  acute  yellow 
atrophy  is  characterized  by  an  early  necrosis  of  a  large  proportion 
of  the  liver-cells,  the  nucleus  becoming  unstainable  while  the  cyto- 
plasm is  still  little  altered  in  appearance,  and  fatty  changes  play  a 
subordinate  role  or  are  absent.  As  Anchiitz  says,  the  poison  seems 
to  strike  at  the  life  of  the  cell,  its  nucleus,  while  phosphorus  attacks 
the  cytoplasm.  Furthermore,  the  poison  of  yellow  atrophy  seems  to 
be  very  specific,  for  it  attacks  the  other  organs  of  the  body  almost 
not  at  all,  and  within  the  liver  it  affects  only  the  hepatic  cells  proper, 
while  the  bile-duct  epithelium  and  the  stroma  cells  are  so  little  in- 
jured that  they  are  able  to  proliferate  greatly,  this  proliferation 
being  a  prominent  feature.  There  are  also  clinical  and  chemical  dif- 
ferences that  will  be  discussed  later,  but  yet,  on  the  whole,  the  re- 
semblances of  yellow  atrophy  and  phospliorus  poisoning  are  so  great 
that  we  have  obtained  much  information  concerning  the  former  by 
means  of  experimental  studies  of  phosphorus  poisoning. 

Delayed  Chloroform  Poisoning. — After  chloroform  narcosis,  and 
rarely  after  etlier,  there  occasionally  develops  a  severe  intoxication, 
with  clinical  and  anatomical  findings  very  similar  to  acute  yelloAV 
atrophy  and  phosj)horus  poisoning;""  in  point  of  the  fatty  changes 
the  cases  usually  stand  intermediate  between  acute  yellow  atro]ili,\'  wnd 
phosphorus  poisoning.     Tliis  action  of  chloroform  would  seem,   fioni 

75  See  Anschiitz,  Arb.  a.  <1.  I'atli.  hist.  Tiiljiiiucn.  1!M>2  ( :! ) ,  -I'-W:  raltaiif.  \vy\\. 
Deut.  Path.  Gesell.,  VMY.\  (5),  !)1:  liicss,  ]?('rl.  klin.  WO.li.,  litOf)  (42).  No.  44a, 
p.  54. 

7«  Complete  review  and  lileratvire  bv  Hevan  ami  Favill.  .Tour.  Amer.  Med. 
Assoc.,  100,5  (45),  091;  :Muskeiis.  Mitt,  (irenz.  :\led.  u.  (hir.,  IDll  (22),  .lliS.  Full 
dineuKsion  of  clieniislry  of  clilorofonu  necrosis  liy  Wells,  .lour.  Riol.  Cheni..  1!>()S 
(5),  12!>.  Kxpciimcnial  necrosis — see  \\'liip))le  and  Sperrv,  .lohns  Hopkins 
Hosp.  Bull.,  I'JU!)    (20),  278;  (Jraiuim,  .lour.  Ivxper    Med.,  1!)12    (15),  '^Ol. 


ACITK   YELLOW    ATh'OI'll)    <)l'  Tin:   Ll\  El!  541 

the  studies  of  Evarts  Graham,'"-'  to  be  produced  by  the  hydrochloric 
acid  formed  from  it  in  the  liver.  Some  cases  of  puerperal  eclampsia 
also  i)resent  such  profound  liver  changes  that  they  are  distinguished 
as  eclampsia  chictiy  on  the  basis  of  the  convulsive  manifestations, 
rather  than  on  the  ground  of  anatomical  changes.  So,  too,  the  hepa- 
tic changes  in  certain  septicemias  and  acute  syphilis  may  resemble 
those  of  acute  yellow  atrophy  to  a  greater  or  less  degree. 

Summary  of  Views  on  Etiology. — From  a  review  of  the  literature 
and  the  study  of  a  few  cases,  the  writer  has  reached  the  following 
understanding  of  the  condition  described  as  acute  yellow  atrophy  of 
the  liver:  The  "atrophy"  is  due  entireh'  to  autolysis  of  necrotic 
liver-cells  by  their  own  enzymes.  In  the  most  typical  cases  of  "pri- 
mary'' or  "idiopathic"'  yellow  atrophy  we  have  to  do  with  a  poison 
having  a  very  specific  effect  on  the  liver-cells,  which  destroys  their 
"life"  (i.  e.,  stops  synthetic  activities)  without  injuring  their  intra- 
cellular proteolytic  enzymes,"  and  consequently  autolysis  occurs;  as 
the  poison  affects  other  organs  but  little,  the  necrosis  and  autolysis 
continue  until  there  is  so  much  loss  of  liver  function  that  systemic 
poisoning  results  from  the  hepatic  insufficiency  and  from  the  result- 
ing accumulation  of  poisonous  products  of  incomplete  metabolism. 
Tliat  the  intoxication  comes  in  large  measure  from  the  changes  in 
the  liver,  even  in  phosphorus  poisoning,  is  shown  by  the  greater  re- 
sistance to  phosphorus  of  dogs  with  Eck's  fistulas."*  The  patient 
dies  from  this  poisoning,'^  and  the  liver  is  found  at  autopsy  to  have 
decreased  by  from  one-third  to  one-half  or  more  in  its  volume.  This 
great  change  would  not  be  possible  if  the  poisons  affected  the  heart, 
kidneys,  or  brain  as  much  as  they  do  the  liver  structure,  which  is 
probabl}'  the  reason  that  phosphorus,  bacterial  poisons,  snake  poisons, 
and  other  poisons  that  destroy  liver-cells  do  not  ordinarily  produce 
the  typical  picture  of  liver  atrophy.  When  these  poisons  affect  the 
liver  more  and  the  other  tissues  less,  we  approach  the  condition  of 
acute  yellow  atrophy;  e.  g.,  if  the  dose  of  phosphorus  is  not  so  great 
as  to  kill  the  patient  through  injury-  of  other  more  vital  organs, 
after  a  few  days  the  necrosed  liver-cells  undergo  autolysis,  and  if 
enough  liver-cells  have  been  destroj-ed,  hepatic  insufficiency  may 
cause  death,  with  the  finding  of  an  anatomical  condition  in  the  liver 
that  can  be  properly  designated  as  acute  atrophy.  Hence  it  is  pos- 
sible for  many  poisons  to  cause  this  condition  under  certain  circum- 
stances, and  there  seem  to  be  certain  unknown  poisons  (probably  of 

76a  .Jour.  Exp.  Med.,  1015    (22),  48. 

~~  According  to  some  investigators  phosphorus  augments  autolysis  even  in  vitro 
(see  Krontowski,  Zeit.  f.  Biol.";  IfllO    (54) ,  479) . 

T8  Fischler  and  Bardach.  Zeit.  pliysiol.  Chem..  1912  (78).  4:^5. 

79  The  mortality  of  cases  sutticiently  typical  to  be  diagnosed  antemortem  is 
estimated  by  Rondaky  (Roussky  Vratch.'Oct.  28.  1900)  at  97  to  98  per  cent. 
Concerning  the  regenerative  changes  in  the  cases  which  recover,  see  Yamasaki 
(Zeit.  f.  Heilk.,  Path.  Abt.,  1903   (24),  248). 


542  ABXOiniMJTIES    l.\    METABOlAfiM 

intestinal  origin  -")  that  are  of  suck  a  iiature  that  they  cause  spe- 
cifically acute  hepatic  atrophy.  The  above  hypothesis  seems  to  ex- 
plain all  the  known  facts  concerning  this  disease.  That  phosphorus, 
chloroform,  and  some  other  poisons  lead  particularly  to  fatty  changes 
may,  perhaps,  be  due  to  tlieir  acting  especially  upon  the  oxidizing  en- 
zymes,*^ leaving  the  autolytic  enzymes  and  the  lipase  free  to  digest 
the  cell  and  to  form  fat.^-  That  it  is  particularly  the  oxidizing  en- 
zymes that  are  attacked  is  well  shown  by  the  chemical  findings,  and 
also  by  Loewy's^^  observation  that  in  poisoning  with  CNH,  which 
acts  by  impairing  oxidation,  the  alterations  in  protein  metabolism  are 
ver}'  similar  to  those  of  phosphonis  poisoning.^*  To  be  sure,  Lusk  ^^ 
found  no  deficiency  in  general  oxidation  in  phosphonis  poisoning, 
but  this  does  not  signify  that  tlie  local  changes  do  not  depend  upon 
local  defective  oxidative  processes. 

Not  only  phosphorus  but  many  metals,  especially  mercury,  seem 
able  to  cause  the  anatomical  changes  of  acute  yellow  atrophy,  for  the 
condition  has  been  observed  very  frequently  in  persons  receiving 
mercurial  and  arsenical  treatment  for  syphilis.^*'  Here  the  syphilis 
has  been  held  responsible  hj  some,  but  the  fact  that  in  many  of  the 
eases  the  syphilis  was  quiescent  or  chronic  at  the  time,  and  that  mer- 
cury and  arsenic  are  known  to  kill  cells  and  stimulate  autolj'sis,  seems 
to  incriminate  the  metals,^'  at  least  in  some  cases. 

CHEMICAL  CHANGES  OF  ACUTE  YELLOW  ATROPHY 

The  Urine. — Most  striking,  and  long  regarded  as  pathognomonic, 
is  the  presence  of  leucine  and  tyrosine  in  the  urine,  first  described 
by  Frerichs.  While  we  now  know  that  these  and  other  amino-acids 
may  occur  in  the  urine  in  any  conditions  in  which  there  is  a  great 
breaking  down  of  tissue  within  the  body,  yet  it  is  true  that  in  no 
other  condition  are  they  found  so  abundantly  as  in  acute  hepatic 
atrophy  (as  high  as  1.5  gm.  of  tyrosine  per  diem  has  been  found). ®^ 

80  See  Carbone,  Riforma  Med.,  1902    (1),  687  and  608. 

81  See  Verworn,  Dout.  med.  Woch.,  1009  (35),  1593;  Joannovics  and  Pick, 
Arch.  fjes.  Physiol.,   1911    (140),  327. 

82  Wells,  Jour.  Amer.  Mod.  Assoc,   1006    (46),  341. 
S3  Cent.  f.  Physiol.,   1906    (19),  23. 

8-t  The  liypothosis  siiofffcstod  by  Quincke  (Xothnatrel's  Handbook,  1899.  vol.  18, 
p.  307)  that  possibly  rofjurfjitation  of  pancreatic  juice  up  tlie  bile  ducts  mipht 
lie  responsible  for  the  dopenerative  chanfjes  in  the  liver,  is  contradicted  by  the 
fact  tliat  the  bile  pressure  is  greater  tlian  tiie  pancreatic  juice  pressure,  and  that 
the  bile-ducts  and  periplieral  portions  of  tlie  h>bules  are  least  afTected.  Nor 
could  Best  "0  prove  that  trypsin  injected  int^  tlie  liver  by  way  of  the  bile-duets  is 
able  to  cause  such  c]ian<res.      (Soo  Wells  and  Passoe.'-J) 

85  Science  of  Nutrition.   Philadelphia,    1909. 

SfiSeverin,  Zcit.  klin.  Mod.,  1912  (76),  138.  Pendip.  :Miinch.  med.  Wooli..  191.-) 
(62),   1144. 

STTileston  (Boston  :\Io(l.  and  Surrr.  Jour..  1908  (158).  510)  lias  described  a 
case  of  acut<»  yellow  atrojihy  from  Tuercurialism  without  syphilis. 

88  An  interostinp  exccjition  has  boon  reported  liy  W.  0.  Smith  (Practitioner, 
1903  (70),  155)  who  found  proat  quantities  of  leucine  in  the  urine  of  a  younp 
woman  who  was  apparently  not  at  all  ill.     Rosenbloom  has  found  tyrosine  crys- 


ACUTE  YEIJ.OW  M'linl'IIY  or  Till:   LIVER  543 

They  arc  iiearl}-  constantly  present  (in  thirteen  out  of  fourteen  cases 
studied  by  Riess),***  tyrosine  being  usually  the  more  abundant. 
Deutcro-proteose  is  also  frequently  (but  not  constantly)  found,  as 
further  evidence  of  abnormal  protein  splitting.'"^  Uric  acid  and 
other  purines  are  often  somewhat,  but  not  characteristically,  in- 
creased, probabl}-  resulting  from  the  nuclear  destruction  in  the  liver. 
There  is  often  an  increase  in  ethereal  sulphates  (Salkowski),"^  and 
in  })h()sp]i()rus  poisoning  various  bases  have  been  found  in  the  urine,"- 
Avhich  i)resumably  might  also  be  found  in  acute  yellow  atrophy  if 
sought  for.  The  total  elimination  of  nitrogen  is  increased  °^  (par- 
ticularly if  the  scanty  intake  is  considered),  and  the  proportion  that 
appears  as  urea  is  decreased,  largely  because  of  the  presence  of  much 
ammonia,"*  part  of  which,  at  least,  is  eliminated  combined  with  or- 
ganic acids.  Chief  of  these  acids  is  sarcolactic  acid,  but  of  partic- 
ular interest  is  the  supposed  appearance  of  oxymandelic  acid, 

HO  /      \  CHOH— COOH, 

which  might  be  derived  from  tyrosine  (Schultzen  and  Ries), 

HO  <(^    ^  Cn,— CH  ( XH, )  —COOH, 

by  the  splitting  out  of  the  NHo  group,  the  benzene  nucleus  failing 
to  be  completely  oxidized,  as  it  normally  is.  The  researches  of  El- 
linger  and  Kotake,"^  however,  make  it  probable  that  the  supposed 
oxymandelic  acid  is  something  else,  most  likely  p-oxyphenyl-lactic 
acid, 


HO  <  >  CH„  —  CHOH  —  COOH 


which  can  be  demonstrated  in  the  urine  of  dogs  poisoned  with 
phosphorus,  and  which  represents  a  simple  deaminization  of  tyrosine 
without  further  oxidation.  It  is  evident  from  the  urinary  findings, 
therefore,  that  oxidation  is  decreased,  which  is  presumably  because 
of  the  destruction  of  liver  tissue  with  its  important  oxidizing  func- 

tals  in  the  urine  of  a  healthy  precmant  woman,  and  cites  other  cases  of  tyrosin 
uria  witliout  hepatic  atrophy   (X.  Y.  ^led.  Jour.,  Sept.  1!),  1914). 

80  Rerl.  klin.  Woch.,  1905  '(42),  Xo.  44  a.,  p.  54. 

90  Salkowski  (Berl.  klin.  Woch.,  inO,5  (42),  15S1 )  found  in  the  urine  of  a  case 
of  acute  yellow  atrophy  a  large  quantity  of  nitrogen  in  a  colloidal  but  non- 
protein form,  apparently  of  carbohydrate  nature.  Mancini  (Arch,  di  farm, 
sperim.,  1906,  Bd.  v)  also  observed  an  increase  in  the  colloidal  nitrogen  of  the 
urine  in  liver  diseases. 

oiVirchow's   Arch.,   1909    (198),   188. 

92Takeda.  Pfliigcr's  Arch.,  1910   (133),  365. 

93  See  Welsch,  Arch.  int.  pharm.  et  th^r.,  1905    (14),  211. 

94  See  Voegtlin,  .Johns  Hopkins  Hosp.  Bull.,  1908  (19),  50:  White,  Boston 
Med.  and  Surg.  .Jour.,  1908    (158),  729. 

95Zeit.  physiol.  Chem.,  1910  (65),  397  and  402;  also  Fromherz,  ibid..  1911  (70), 
351. 


544  AB\OR.UALITJi:S    IN    METABOLISM 

tions.  The  reduction  of  oxidation  eau  also  be  shown  experimentally 
by  studying  the  respiratory  exchange,  Welseh  having  found  the  oxi- 
dation decreased  by  from  %  to  Y:,  in  phosphorus  poisoning.  Carba- 
mates do  not  seem  to  be  present  in  reeogni/.al)le  amounts,  and  sugar 
is  also  absent. 

In  phosphorus  poisoning  the  urinary  findings  are  similar,  but  with 
marked  (pumtitative  differences.  Tyrosine  cannot  usually  be  de- 
tected, at  least  by  ordinary  methods,  being  found  by  Riess  in  but  7 
of  36  cases  of  (human)  phosphorus  poisoning,  and  in  but  4  of  these 
was  it  abundant.  Leucine  is  even  less  frequently  found.  With  ex- 
perimental animals  glycocoll  and  other  amino-acids  have  been  found  ^ 
in  the  urine,  and  they  could  probably  be  found  in  acute  hepatic  atro- 
phy if  the  same  delicate  methods  were  employed.  AVohlgemuth  -  has 
indeed  found  glycocoll,  alanine,  and  arginine  in  human  urine  after 
phosphorus  poisoning.  The  small  quantity  of  amino-acids  in  phos- 
phorus poisoning  is  probably  due  to  the  relative  slowness  of  the  auto- 
lytic  changes.  On  the  other  hand,  the  deficiency  of  oxidation  in  phos- 
phorus poisoning  is  shown  by  the  abundant  elimination  of  organic 
acids,  Riess  having  obtained  as  high  as  4  to  6  grams  of  the  zinc  salt  of 
paralactic  acid  from  the  urine  (per  liter)  in  human  cases,  and  its 
presence  seems  to  be  constant. 

The  Liver.^ — In  the  liver  may  be  found  an  abundance  of  the  free 
amino-acids  that  have  not  yet  escaped  by  diffusion,  their  presence 
having  been  first  detected  by  Frerichs  microscopically.  Taylor  *  was 
able  to  isolate  from  a  liver  weighing  900  grams,  0.35  gm.  of  leucine 
and  0.612  gm.  aspartic  acid,  which  probably  represent  much  less  than 
the  total  amount  present.  Deuteroalbumose  was  also  found,  but  no 
peptone,  arginine,  histidine,  or  lysine,  and  glj^cogen  was  also  absent. 
In  another  case  that  appeared  to  be  the  result  of  chloroform  intoxi- 
cation, Taylor  ^  obtained  4  grams  of  leucine,  2.2  grams  of  tyrosine, 
and  2.3  grams  of  arginine  nitrate.  AVells  found  several  amino-acids 
free  in  sufificient  quantity  to  identify  in  the  liver  in  cases  of  acute  yel- 
low atrophy  and  chloroform  necrosis,  an  increase  in  gelatigenous 
substance  in  tlie  former,  and  of  organic  non-lipoidal  phosphorus  in 
both,  sulphur  being  unchanged.  The  increase  in  tissue  phosphorus 
is  striking,  and  agrees  with  Slowtzoff's  and  Wohlgemuth 's "  finding 
that  the  tissue  phosphorus  persists  in  experimental  phosphorus  poi- 
soning.    Wakeman  '  found  that  in  phospliorus  poisoning  of  dogs  the 

1  Abderhalclen  and  Barker.  Zcit.  plivsiol.  Cliom..  l!)04  (42),  524;  AlKlorliald.-n 
and  Bcrgcll,  ibid.,   li)()3    CM),  4G4. 

^  Zeit.   physiol.  Cliem.,    1!)(),5    (44),   74. 

3  Full  analyses  and  discussion  of  the  clieinistry  of  the  liver  in  acute  yellow 
atrophv  and  clioloroforni  necrosis  <;iven  hv  Wells.  Jour.  Kxper.  iled.,  11I07  (H), 
627;  Arch.  Jnt.  .Med.,  litOS   (1),  .-)S!t:  .lour."  Biol.  Cheni.,  l!tOS   ( f) ) .  12!). 

4  Zeit.  pliysiol.  Ciieni.,  I!t(t2   (;i4).  r,A{);  .lour.  .Med.  Research.  1!)02   (8).  424. 

5  Tniv.  of  Calif.  Pui)lications   (Pathol.),  l'.»04    (1),  4.S. 
'•  Biodiem.  Zeit..  1011    (.S2),  172. 

-Jour.  Kxper.  Med.,  1!)05    (7),  292;   Jour.  Jiiol.  V\wm.,   lUUS    (4),  11!). 


ACUTE  YELLOW  ATROPHY  OF  THE  LI]  Eli  545 

liver  shows  a  diniimitioii  of  the  hexone  bases  as  a  whole,  the  arginiue 
being  especially  reduced;  but  no  such  change  was  found  by  him  in 
acute  yellow  atrophy,  nor  by  Wells  in  chloroform  necrosis.  Jack- 
son and  Pearce  ^  found  an  increase  in  the  diaraino  nitrogen  with  ex- 
tensive necrosis  of  the  liver  in  dogs  and  liorses.  Wohlgemuth  '■*  found 
arginine  in  the  urine  in  phosphoiiis  poisoning.  The  lecithin  of  the 
liver  is  also  decreased  (Heffter^°  and  Wells),  and  the  increase  in 
P0O3  observed  in  the  urine  presumably  comes  partly  from  this  source ; 
cholesterol  is  unchanged.  Beebe  ^^  found  the  pentose  of  the  liver  not 
greatly  altered  from  the  normal  relations.  The  typical  idiopathic 
atrophied  liver  shows  little  or  no  inorease  in  fat,  either  chemically  or 
microscopically,  whereas  there  is  considerable  replacement  of  the  lost 
liver  substance  by  water,  as  shown  in  the  following  table: 

Fat-free 
Dried 
"Water  Fat  Substance 

Normal   liver    (Quincke)     76.1  3.0  20.9 

Normal   liver    (Wells)    77.6  5.0  17.4 

Acute  atrophy   (Perls)    81.6  8.7  9.7 

Acute  atrophy    (Perls)     76.0  7.6  15.5 

Acute  atrophy    (v.  Starck)    80.5  4.2  15.5 

Acute  atrophy    (Taylor)     85.8  2.0  12.2 

Acute  atrophv    (Wakeman)     79.3  .   .  .     . 

Acute  atrophy    (Wells)     83.8  2.5  13.7 

Acute  atrophy   (Voegtlin) 78.0  6.6  15.4 

Phosphorus  poisoning    (v.   Starck)     60.0  29.8  10.0 

Fatty   degeneration    (v.    Starck)     64.0  25.0  11.0 

Chloroform  necrosis    (Wells)    72.4  8.8  18.8 

Similar  results  have  been  obtained  frequently  by  other  observers.  Tay- 
lor estimating  that  in  his  ease  about  three-fourths  of  the  liver  paren- 
chyma had  disappeared.  The  yellow  color  of  the  liver  tissue  charac- 
teristic of  this  condition  seems  to  be  due  to  bilirubin  rather  than  to 
fat,  because  as  soon  as  the  tissues  are  put  into  oxidizing  agents  {e.  g., 
dichromate  hardening  fluids)  they  turn  grass-green  from  the  oxida- 
tion of  the  bilirubin  into  biliverdin.  There  seems  to  be  a  marked  in- 
crease in  free  fatty  acids,  probably  the  unsaturated  higher  fatty 
acids,  which  are  strongly  hemolytic.^" 

Jacoby  ^^  found  that  the  livers  from  phosphorus-poisoned  dogs 
underwent  autolysis  with  greater  rapidity  than  normal  livers,  which 
was  attributed  to  increased  activity  or  amount  of  the  autolytic  en- 
zymes, although  addition  of  phosphorus  to  a  solution  containing  liver 
feraients  was  not  found  to  increase  their  activity.  The  aldehydase 
was  not  found  decreased,  and  tyrosinase  could  not  be  demonstrated, 

s  Jour.  Exper.  Med.,  1907   (9),  520. 

9  Zeit.  physiol.  Chem.,  1905    (44),  74. 

10  Arch.  exp.  Path.  u.  Pharm.,   1891    (28),  97. 

11  Amer.  Jour,  of  Physiol.,   1905    (14),  237. 
i2Joannovics  and  Pick,  Berl.  klin.  Woch.,   1910    (47),   928. 

13  Zeit.   physiol.   Chem..   1900    (30),    174;    see  also   Porges  and  Pribram,   Arch, 
exp.  Path.  u.  Pharm.,   1908    (59),  20. 
35 


546  Aii\oiniMJTii:s  /.v   metauolism 

but  SloAvtzoff  ^*  found  both  peroxidase  and  protease  decreased,  and 
iitti-ibutcd  tlie  increased  autolysis  to  a  g'reater  acidity  of  the  liver. 

The  Blood. — In  the  blood  marked  changes  are  found,  one  of  the 
most  prominent,  besides  the  icterus,  being  the  decreased  coagulability 
of  the  blood.  This  seems  due  to  a  loss  of  fibrinogen, ^^  wliich,  with 
the  giobuliu,  is  greatly  decreased,  the  albumin  remaining  less  al- 
tered.^® The  fibrin-ferment  also  seems  to  be  decreased.  These 
changes  may  be  due  to  direct  autolysis  of  the  blood  constituents  (Ja- 
coby  having  found  that  thrombi  become  rapidly  dissolved  in  phos- 
pliorus-poisoning)  or  to  the  changes  in  the  liver.  The  icterus  de- 
pends apparently  upon  lesions  of  the  finest  bile  capillaries,^^  although 
there  is  also  some  increase  in  hemolj-sis,  and  a  decrease  in  the  total 
blood  and  all  its  elements  (Welsch)  ;  ^^  and  both  bile  salts  and  pig- 
ments appear  in  the  urine.  In  all  these  diseases  with  marked  liver 
changes  there  is  an  increase  in  the  lipase  of  the  blood. ^^^  Neuberg 
and  Richter  ^^  have  analyzed  the  blood  drawn  during  life  from  a  pa- 
tient with  acute  hepatic  atrophy,  and  isolated  from  355  c.c.  of  blood 
0.787  gm.  tyrosine,  1.102  gm.  leucine,  and  0.240  gm.  of  lysine ;  they 
estimated  the  amount  of  free  amino-acids  in  the  entire  blood  to  be 
about  30  grams.  This  amount  is  so  large  that  they  question  the  pos- 
sibility of  it  all  arising  from  the  degenerated  liver  tissue ;  but  more 
analyses  are  necessary  before  conclusions  on  this  point  can  be  drawn,-*' 
especially  by  the  use  of  the  newer  methods.  Certainly  in  dogs  suf- 
fering from  ehlorofomi  necrosis  of  the  liver  or  phosphorus  poisoning 
the  amount  of  free  amino  acids  in  the  blood  and  urine  is  usually  very 
small. -"•■' 

Origin  of  the  Amino=acids. — The  earliest  conception  of  the  source 
of  the  leucine  and  tyrosine  found  in  the  urine  was  that  it  came  from 
the  products  of  tryptic  digestion  absorbed  from  the  intestinal  tract, 
which  the  liver  could  not  convert  into  urea  because  of  its  damaged 
condition.     On  the  demonstration  by  Jacoby  -^  that  these  same  bodies 

i4Biocliom.  Zcit.,   mil    (31).  227. 

1^' Whipple  and  ITurwitz  (Jour.  Exper.  IVled.,  1011  (l:^).  i:}(i)  find  a  <,noat 
(IccToase  in  fi1)rinof;on  during  experimental  cliloroforin  necrosis  of  the  liver. 

i''.Jacobv,  lor  cit.;  see  also  Doyon,  C'ompt.  Rend.  Soc.  Biol.,  1!M(.")  (.IS).  403; 
and  1900,  Vol.  06. 

1"  Lang  (Zoit.  exp.  Path.,  1006  (3),  473)  found  fihrinogen  in  the  liih"  of  a 
dog  poisoned  with  phospliorus,  which  may  account  for  the  oeel\isien  of  the 
bile  vessels  and  tlie   resulting  jaundice. 

18  Arch.  int.  Pharm.  et  Ther.',  lOO.")    (14),  107. 

i«a  Whipple  et  al..  Pull.  Johns  Hopkins  ITosp.,  1013  (24),  207  and  357. 
Quinan  found  the  lipase  content  of  liver  tissue  much  reduced  in  chlorofium 
necrosis  (.Tour.  ^led.  Res.,  1015  (32),  73).  A  review  of  work-  |)uhlished  on  blood 
dianges  and  liver  fuTiction  in  phosphoiMis  and  cliloroforin  ]ioisoninL;  is  <^iveii  by 
^^arshalI  and   liowntree,  Jour.  Exp.  Med..   1015    (22),  33;i. 

ii'Dcut.med.  Woch.,  1004    (30),  400. 

20  V.  Bergmann  (llofnieister's  l^eit.,  1004  (6),  40)  was  able  to  isolate  2.3  grams 
of  amino-acids  combined  with  the  chloride  of  naphthalene  sulplioiiic  arid,  from 
270  c.c.  of  blood  in  a  case  of  acute  vellow  atrophv. 

■iOaSee  Van  Slvke,  Arch.  Int.  IMed.,  1017    (10),"  77. 

21  Zeit.  physiol.  Chcm.,   1000    (30),   174. 


ACID  JSTOXICATION  547 

were  present  in  the  livers  of  phosphorus-poisoned  animals  because  of 
autolysis,  it  became  probable  that  the  leucine  and  tyrosine  found  in 
the  urine  were  formed  from  the  degenerated  liver-cells  rather  than 
in  the  intestine,  which  view  has  become  generally  accepted.  It  seems 
most  probable,  however,  that  the  urinarj^  amino-acids  are  derived 
parti}'  (and  perhaps  chiefly)  from  the  autolysis  of  the  liver,  and 
partly  from  amino-acids  produced  both  in  the  intestine  and  within 
the  body  during  tissue  metabolism,  and  which  the  liver  cannot  trans- 
form into  urea  as  it  normally  does,  for  several  observers  have  re- 
ported that  even  relatively  slight  disturbances  in  hepatic  function 
are  accompanied  by  a  considerable  rise  in  the  amino-acids  in  the 
urine.^- 

ACID  INTOXICATION -3 

If  a  rabbit  is  given  in  repeated  small  doses  by  mouth  considerable 
quantities  of  inorganic  acids,  such  as  hydrochloric  or  phosphoric  acids, 
which  it  cannot  destroy  by  oxidation,  it  soon  becomes  extremely  ill. 
The  manifestations  are  characteristic — unsteadiness  of  motion  and 
stupor  being  followed  by  coma,  in  which  the  striking  feature  is  the 
excessively  active  respiration,  as  if  the  animal  were  being  asphyxi- 
ated (the  so-called  "air  hunger"),  while  at  the  same  time  there  is  no 
cyanosis  and  the  blood  is  bright  red,  containing  much  less  COo  than 
normal,  while  the  amount  of  oxygen  remains  quite  normal.  The  cur- 
rent explanation  of  this  interesting  condition  is  as  follows :  Normall}^ 
the  blood  carries  the  C0„  away  from  the  tissues  to  the  lungs  in  com- 
bination with  the  inorganic  alkalies  of  the  blood,  of  which  sodium  is 
by  *far  the  most  abundant.  This  combination  is  the  bicarbonate  of 
sodium  for  other  base),  which  in  the  lungs  is  decomposed  into  the 
carbonate,  the  COo  escaping  into  the  alveolar  air,  according  to  this 
equation : 

2X:iHCO,^Xa,C0,  -f  H,0  +  CO, 

The  carbonate  thus  formed  goes  back  to  the  tissues  to  combine  again 
with  more  COo  and  form  bicarbonate.  If  acids  are  introduced  into 
the  blood  they  combine  with  the  alkalies  there,  forming  neutral  salts 
which  are  eliminated  in  the  urine,  and  in  this  way  the  amount  of 
alkali  in  the  blood  is  reduced,  with  a  consequent  reduction  in  the  ca- 
pacity of  the  blood  to  carr\^  CO.,  away  from  the  tissues;  the  amount 
of  COo  in  the  blood  sinking  to  as  low  as  2.5  and  3  per  cent.  (Walter). 
Consequently,  in  acid  poisoning  the  CO,  produced  in  metabolism  ac- 
cumulates in  the  tissues  where  it  is  formed,  and  blocks  the  processes 

22  See.  Masuda.  Zeit.  exp.  Path.,  1911  (S),  r>20:  Labhe  and  Bitli.  Compt.  Rend. 
Soc.  Biol..  1012    (73),  210. 

23  General  literature  to  lOOS,  piven  by  Ewing,  Arcli.  Tnt.  "Nfed..  inns  (2),  .3.30: 
also  see  Magnus-Levy,  Ergebnisse  inn.  Med.,  1008  (1),  374:  Lusk,  Arch.  Int. 
Med.,  1900  (3),  1.  More  recent  literature  given  by  PTurtley.  Quart.  ,Tour.  ]\Ted., 
1016  (9),  301,  and  an  excellent  review  of  recent  work  bv  ^^^litnev,  Bost.  ^led. 
Surg.  Jour.,  1017    (176),  22-5. 


548  ABNORMALITIES    IX    METABOLISM 

of  oxidation,  so  that  tlie  animal  suffers  from  asphyxia  exactly  as  if  it 
were  deprived  of  air.  In  other  Avords,  the  lack  of  alkalies  in  the 
blood  in  acid  intoxication  checks  the  "internal  respiration,"  as  in- 
tracellular gas  exchange  is  called,  by  preventing  the  removal  of  COg 
from  the  cells.  The  acids  stimulate  the  respiratory  center,  which  is 
extremely  sensitive  to  them,  and  the  increased  respiration  tends  to 
reduce  the  acidity  by  getting  rid  of  the  CO,,-*  but  under  the  condi- 
tions of  the  experiment  this  is  not  sufficient  to  prevent  asphyxia. 

If  the  urine  of  such  an  animal  is  analyzed,  it  is  found  to  contain 
increased  quantities  of  the  four  chief  inorganic  bases,  Na,  K,  Ca,  and 
Mg  (the  last  two  apparently  being  derived  from  the  bones),  but  in 
addition  to  these  it  is  found  that  the  amount  of  ammonia  in  the  urine 
is  decidedly  increased.  If  instead  of  a  rabbit  a  carnivorous  animal, 
such  as  a  dog,  is  given  acids,  it  will  be  found  relatively  insusceptible, 
so  that  great  quantities  can  be  given  without  causing  acid  intoxica- 
tion. Examination  of  the  urine  of  such  a  dog  will  show  that  the 
elimination  of  ammonia  is  increased  much  more  than  it  is  in  the 
herbivora,  while  the  inorganic  alkalies  are  increased  but  little.  From 
this  it  is  deduced  that  in  acid  intoxication  part  of  the  nitrogen  that 
normally  goes  to  form  urea  becomes,  while  in  the  antecedent  form  of 
ammonia,  combined  with  part  of  the  acid  that  has  entered  the  blood. 
In  this  way  much  of  the  neutralization  of  the  acids  is  accomplished 
by  ammonia,  and  the  inorganic  alkalies  of  the  blood  are  spared. 
As  in  carnivora  the  amount  of  protein  metabolism  is  much  greater 
and  more  rapid  than  in  herbivora,  the  ammonia  available  for  neutral- 
ization of  acids  is  much- greater  than  in  the  latter,  and  hence  the  rela- 
tive lack  of  susceptibility  of  carnivora  to  acid  poisoning.-^  Accord- 
ing to  Landau,-*'  the  proteins  of  the  blood  also  combine  much  of  the 
acid.  Another  factor,  which  is  commonly  overlooked,  is  the  possible 
accumulation  of  acids  within  the  cells,  which  must  modify  greatly  any 
conclusions  based  upon  studies  of  the  blood  and  urine.  It  is  witliin 
the  cells  that  the  effects  of  acids  must  be  manifested,  and  it  is  per- 
fectly possible,  and  indeed  almost  certain,  that  we  may  have  degrees 
of  acidity  and  alkalinity  in  the  cells  which  are  quite  different  from 
those  in  the  blood. 

As  pointed  out  especially  by  Henderson,-"'^  the  normal  reaction  of 
the  body  is  kept  practically  constant  chietiy  by :  1.  The  salts  of  COo 
and  H3PO4,  existing  in  such  proportions  of  carbonate,  bicarbonate 
and    carbonic    acid,    or    disodium-    and    monosodinm-hydrogen-phos- 

24  Sop  Porpos,  Wion.  klin.  Wooli..  1011    (24).  1147. 

2r.  Tliis  lias  Ih'oti  iiiooly  sliown  liy  Kppin},'6r  (\\ii>ii.  klin.  Wooh..  1006  (10).  111). 
who  found  tliat  administration  of  considorablo  quantities  of  amino-acids  (plyco- 
ooll,  alanine,  aspartic  acid)  to  rabbits  proatly  inoroasod  tlieir  resistance  to  acid 
intoxication,  presunialdy  by  yielding  ammonia  tlirougli  normal  steps  of  pro- 
tein metabolism. 

2<!  Arch.  exp.  Path.  u.  Pharm.,  1000   (52),  271. 

2«a  See  Tlarvej'  Society  Lectures,   1014-15. 


ACID  JXrOXICATIOX  549 

phate,  as  to  produce  an  almost  neutral  solution.  These  being  salts  of 
weak  acids  with  strong  bases  it  follows  that  when  a  stronger  acid, 
such  as  lactic  or  butyric  combines  with  the  bases  there  is  only  the 
weak  acid  liberated,  and  hence  the  influence  of  the  strong  acid  on  the 
blood  reaction  is  greatly  reduced.  (2)  The  acid  most  abundantly 
formed  in  metabolism,  COo,  is  volatile  and  hence  is  rapidly  excreted 
by  the  lungs  without  withdrawing  bases  from  the  blood.  (3)  The 
kidneys  can  eliminate  the  other  buffer  acid,  PO4,  with  but  a  minimum 
of  base  attached  in  the  form  of  NaH.PO^ ;  and  they  also  remove  the 
basic  product  of  metabolism,  ammonia.  By  the  combined  influence  of 
these  factors  the  acids  formed  in  metabolism  are  passed  out  with  a 
maxinuim  rapidity  and  with  a  minimum  alteration  in  the  reaction  of 
the  fluids  by  which  they  are  carried  through  the  body.  In  addition 
to  these  we  have,  as  mentioned  before,  the  capacity  of  the  proteins  to 
combine  with  both  acids  and  alkalies,  the  reserve  neutralizing  capac- 
ity of  ammonia  found  in  metabolism,  and  also  the  enormous  reserve 
supply  of  bases  in  the  bone  salts.-*"' 

Acidosis,  therefore,  is  a  condition  in  which  the  essential  feature  is 
the  impoverishment  of  the  body  in  available  bases,  whereby,  there  re- 
sults a  decreased  capacity  of  the  tissues  to  get  rid  of  CO,  and  other 
acids  formed  in  their  metabolism.  This  reduction  in  bases  may  be, 
and  most  usually  is,  the  result  of  excessive  production  of  acids,  in 
excreting  which  the  bases  are  eliminated  in  excess,  but  it  may  also 
result  from  deficient  capacity  of  the  kidneys  to  excrete  acids,  since 
the  kidneys  play  an  important  role  in  regulating  acidity.  The  degree 
of  acidosis  may  be  estimated  in  several  ways,  as  follows :  ^^^ 

1.  By  determining  the  COo  content  of  the  blood,  which  must  de- 
crease as  other  acids  increase,  or  the  bases  decrease. 

2.  Direct  estimation  of  the  H-ion  concentration  of  the  blood. 

3.  By  determining  the  amount  of  acid  or  alkali  necessary  to  change 
the  reaction  of  the  blood  to  different  indicators. 

4.  Determination  of  the  COo  tension  of  the  alveolar  air,  this  vary- 
ing directly  with  the  CO2  tension  of  the  arterial  blood. 

5.  The  "alkali  tolerance  test"  of  Sellards,  which  consists  in  ascer- 
taining the  amount  of  sodium  bicarbonate  that  must  be  taken  by 
mouth  in  order  to  produce  an  alkaline  urine. 

6.  Estimation  of  the  amount  of  organic  acids,  H-ion  concentration, 
and  ammonia  content  of  the  urine ;  a  method  which  is  fundamentally 
defective  since  it  indicates  merely  the  acids  and  bases  that  have  been 
removed  from  the  body  and  not  those  that  remain  to  modify  its  reac- 
tivity. 

7.  Determination  of  the  capacity  of  the  blood  serum  to  bind  COg. 

20b  The  existence  of  the  opposite  condition,  "alkalosis,"  has  not  been  estab- 
lished, unless  it  occurs  in  parathyroid  tetany  (See  Wilson,  Stearns  and  Janney, 
Jour.  Biol.  Chem.,  vols.  21  and  23). 

26c  See  review  by  Frothingham,  Arch.  Int.  Med.  1916   (18)   717. 


550  AUyORMALITIES    IX    METABOLISM 

Normal  serum  binds  about  75  per  cent,  of  its  volume  of  CO,,  whereas 
in  acidosis  it  may  bind  but  20  per  cent.  (Van  Slyke). 

DIABETIC   COMA  23 

In  man,  poisoning  with  inorganic  acids,  as  in  the  experiments  cited 
above,  is  a  rare  occurrence,  but  not  infrequently  acid  intoxication  re- 
sults from  the  presence  of  undue  quantities  of  organic  acids  pro- 
duced in  metabolism.  Tlie  most  striking  example  of  this  is  the  coma 
of  diabetes,  in  which  the  asphyxia  without  cyanosis,  dependent  upon 
failure  of  the  blood  to  carry  CO,,  is  sometimes  strikingly  similar  to 
that  observed  in  experimental  animals  poisoned  with  acids.  In  dia- 
betic coma  the  acid  intoxication  is  due  chiefly  to  the  accumulation  in 
the  blood  or  tissues  of  large  quantities  of  (3-oxyhiityric  acid.  Associ- 
ated with  it,  in  smaller  quantities,  are  usually  found  diacetic  (aceto- 
acetic)  acid  and  acetone,  which  are  chemically  so  closely  related  that 
it  has  been  generally  considered  that  they  are  derived  from  the  oxy- 
butyric  acid,  as  follows : 

y8-oxybutyric  acid  is — 

CH3  — CHOH— CH,— COOH, 

and  by  oxidation  this  readily  forms — 

CH3— CO— CH„— COOH, 

which  is  diacetic  acid   (being  two  molecules  of  acetic  acid  united  to 
each  other,  as  follows)  : 


CH3— CO—  I  OH— H  I  — H,C— COOH. 

Diacetic  acid  is,  in  turn,  readily  deprived  of  its  carbon  dioxide,  forming 
acetone, 

CH3— CO— CH3. 

All  these  reactions  are  easily  accomplished  in  the  laboratory^  and  there 
seemed  to  be  reason  for  believing  that  they  may  normally  occur  in  the 
same  way  in  the  animal  body.  Wakeman  and  Dakin,-'^  and  others, 
however,  found  evidence  that  the  liver  cells  may  also  reduce  diacetic 
to  /8-oxybutyric  acid,  and  it  seems  probable  that  this  is  the  usual 
direction  of  the  reaction,  which  they  have  found  to  be  produced  by  a 
specific  enzyme.  Ilurtley  "^  concludes  that  the  reduction  of  aceto- 
acetic  acid  to  oxybutyric  acid  is  accomplished  b}^  the  body  under 
ordinary  conditions  far  more  readily  than  the  oxidation  of  oxybutyric 
to  aceto-acetic  acid.  Marriot  "^^  gives  the  following  scheme  as  indi- 
cating the  normal  path  of  fatty  acid  catabolism: 

d — oxvhutric    acid 

Fatty  acid  y{  I'.utyric  aci.l(  ?)  ) VAccto-acctic   acid )./< '••^'Klilv  burned) 

1 — oxynutric  acid 
(dillicultly  hurned) 

27  Jour.  Biol.  Chom.,  inon   (6).  373;   1010   (8),  105. 
2-a.T<mr.  Biol.  Chem.,  1914   (18),  241. 


DIABETIC  COMA  551 

The  study  of  tlif  utilization  of  X\ni  acidosis  suljstaiici's  wlicu  injected 
intravenously  at  accurately  measured  rates  for  considerable  periods 
by  AVilder,-"''  furnishes  conclusive  evidence  of  the  origin  of  /3-oxy- 
butyric  acid  from  acetoacetic  acid.  He  found  that  normal  dogs  ex- 
crete ^-oxybutyric  acid  when  it  is  injected  at  the  rate  of  0.4  gm. 
C 0.0032  gm.  molecule)  of  the  sodium  salt  per  kilo  of  body  weight  per 
hour,  but  not  with  lower  rates.  Sodium  acetoacetate  was  excreted 
when  injected  in  rates  of  0.2  gm.  (0.0016  gm.  molecule)  per  kilo  per 
hour,  and  when  injected  at  the  0.4  gm.  rate  it  was  excreted  accom- 
panied by  )8-oxybutyric  acid.  Evidently,  then,  the  acetoacetic  acid 
must  be  converted  almost  quantitatively  into  /8-oxybutyric  acid.  On 
the  other  hand,  acetoacetic  acid  did  not  appear  in  the  urine  when 
larger  quantities  of  /S-oxybutyric  acid  were  injected,  and  hence  it  is 
apparent  that  not  much  if  any  urinary  acetoacetic  acid  is  derived  from 
this  source. 

As  long  as  a  diabetic  is  burning  at  least  one  molecule  of  carbohydrate 
to  three  of  higher  fatty  acids  the  urine  is  free  from  both  acids,  although 
small  amounts  of  actone  (traces  of  which— under  0.02  gm.  per  day 
— occur  in  normal  urine) -^  may  be  present;  but  when  for  any  reason 
the  daily  oxidation  of  carbohydrates  falls  below  this  minimum  the 
two  acids  appear,  combined  largely  with  ammonia,  but  partly  with 
mineral  bases.  Fats  burn  in  the  fire  of  the  carbohydrates  and  as  Wood- 
yatt  ^^*  puts  it,  when  the  proportion  of  fat  is  too  great  for  the  fire 
it  "smokes"  with  unburnt  fats  and  acetone  bodies.  Normally  but  2 
to  5  per  cent,  of  the  nitrogen  of  the  urine  is  in  the  form  of  ammonia, 
but  in  diabetic  acidosis  the  proportion  may  reach  from  10  to  25  per 
cent.,  the  proportion  of  urea  being  correspondingly  reduced.-^ 

The  presence  of  large  quantities  of  these  acids  in  the  urine  presages 
coma,  during  which  the  amount  of  oxybutyric  acid  often  reached  15-20 
grams  per  day,  and  has  been  known  to  reach  150  grams  (Kiilz  claimed 
to  have  found  226  grams).  Diacetic  acid  appears  in  relatively  small 
amounts,  rarely  exceeding  10  per  cent,  of  the  total  organic  acids  of 
the  urine.  When  oxybutyric  acid  is  present  the  other  two  substances 
are  always  present,^"  but  aceto-acetic  acid  and  acetone  are  said  by 
some  to  occur  in  the  absence  of  /^-oxybutyric  acid.  According  to 
certain  observers,  in  the  development  of  acetonuria,  acetone  is  the 
first  of  the  three  bodies  to  appear ;  ^^  when  0.4  te  0.5  gm.  of  acetone  is 

27b  Jour.  Biol.  Chem.,  1917  (.30). 

28  Concernino:  normal  occurrence  of  acetone  in  lilood  and  tissues,  see  Halporn 
and  Landau,  Zeit.  exp.  Path,  u  Ther.,  1906   (3),  4t)0. 

2SaJour.  Amer.  Med.  Assoc.,  1916    (66).   1910. 

29  According  to  Y.die  and  Whitley  (biochemical  -Tour..  1906  (1),  11).  ad- 
ministration of  excessive  amounts  of  alkali  causes,  conversely,  elimination  of 
increased  amounts  of  organic  acids. 

30  See  Pavy,  Lancet,  1902    (ii).  64  et  se</.    (general  review). 

31  Folin  says  tiiat  perfectly  fresh  diabetic  urine  does  not  contain  anv  acetone, 
that  which  is  commonly  found  being  derived  from  diacetic  acid  which  rapidly 
decomposes  into  acetone. 


552  AliXORMA/JTJES    I\    MET  A  HOLISM 

present  iu  the  day 's  urine  diacetic  acid  may  be  found,  but  oxybutyric 
acid  does  not  usually  appear  until  the  amount  of  acetone  exceeds  1 
gram.  After  this  the  chief  increase  is  in  the  oxybutyric  acid,  which 
often  reaches  30  to  80  grams,  whereas  the  diacetic  acid  and  acetone 
together  rarely  exceed  7  to  8  grams  (v.  Noorden).  However,  the 
results  obtained  by  most  investigators  do  not  support  the  regular 
quantitative  relationship  described  above,  extremely  irregular  results 
being  commonly  obtained;  as  a  rule,  when  any  one  of  the  three  ace- 
tone bodies  is  present  in  large  amounts  there  is  an  abundance  of  each 
of  the  others.  Kenneway  ^-  confirms  Neubauer's  statement  that  oxy- 
butyric acid  is  rather  constantly  from  60  to  80  per  cent,  of  the  total 
acetone  bodies  excreted  in  the  urine.  In  the  internal  organs  the  ace- 
tone bodies  may  also  be  detected,  especiallj^  in  the  liver.^^  In  normal 
blood  Marriott  found  less  than  4  mg.  of  oxybutyric  acid,  and  1.5  mg. 
of  acetone  and  aceto-acetic  acid  together,  per  100  c.c. ;  but  in  diabetic 
coma  tlie  figures  rose  as  high  as  45  mg.  and  28  mg.  respectively  for 
eacli  fraction,  the  amount  in  the  blood  not  corresponding  to  the 
urinary  excretion.^^'' 

Relation  of  Acidosis  to  Diabetic  Coma. — Tliere  seems  to  be  lit- 
tle room  for  doubt  that  the  typical  diabetic  coma  with  "air  hunger" 
depends  upon  an  excess  of  these  substances  in  the  blood — i.  e.,  is- 
an  acid  intoxication — for  the  following  reasons:  (1)  The  coma  ap- 
pears when  the  amount  of  organic  acids  in  the  urine  is  highest,  and 
is  absent  when  there  is  little  or  none  of  them  in  the  urine.  (2)  Be- 
cause of  the  resemblance  of  the  symptoms  to  those  of  experimental  acid 
intoxication.  (3)  Because  of  the  repeated  demonstration  of  a  re- 
duced amount  of  alkali  in  the  blood,  as  determined  by  titration,  and 
a  great  reduction  of  the  amount  of  CO,  carried  in  the  venous  blood. 
The  capacity  of  the  serum  to  absorb  CO,  in  vitro  is  also  greatly  re- 
duced, from  a  normal  75  per  cent,  to  as  low  as  20  per  cent.  (Van 
Slyke).  By  means  of  gas-chain  measurements  Roily  ^*  found  that 
by  far  the  lowest  OH  values  ever  observed  in  the  blood  are  in  dia- 
betic coma ;  and  Sellards  ^^  showed  that  in  diabetes  the  tolerance 
for  alkalies  may  be  increased.  (4)  The  marked  improvement  that 
sometimes  results  from  the  administration  of  alkalies  (usually  sodimn 
bicarbonate).  Associated  with  this  improvement  is  an  elimination  of 
greath'-  increased  amounts  of  organic  acids,  indicating  their  previous 
retention  in  the  body  because  of  lack  of  alkali  with  which  they 
could  combine  (or  their  liberation  from  combination  with  proteins — 
Landau).  But  there  are  man.y  cases  of  diabetic  coma  without  typ- 
ical air  hunger,  and  it  is  the  exception  rather  than  the  rule  for  alkali 
therapy  to  produce  a  marked  improvement  in  the   fully   developed 

32Biocliem.  Jour.,   1013    (8),  .•^5.'"). 
33SaRsa,  Bioc'hcm.   Zeit.,   1014    (59).  3()2. 
33a  Marriott,  Jour.  Hiol.  Chem.  1914    (18).  ,'507. 
34Miincli.  mod.  Woeli.,  1912    (."59).  1201. 
SBJolins  iropkins  IIosp.  Bull.,  1912   (23),  289. 


^  RELATlOy  OF  ACIDO.SIS   TO  iJlAJiiyi/C  COMA  553 

coma  of  diabetes.  Fiirtliermore,  coma  may  occur  in  diabetics  who  are 
producing-  no  such  ((uantity  of  oriianic  aeids  as  would  seem  tlieoreti- 
cally  to  be  necessarj^  to  cause  enough  acid  intoxication  to  result  in 
acidosis,  and  coma  develops  in  diabetics  who  are  being  supplied  with 
sufficient  bases  for  all  requirements.  Hence  it  must  be  concluded 
that  only  a  part  of  the  symptomatology  of  diabetic  coma  depends  on 
acids  as  such,  but  as  yet  we  do  not  know  what  other  agents  are 
acting.^" 

/8-ox3'butyric  and  diacetic  acid,  according  to  many  authorities, 
seem  to  have  no  specific  poisonous  effects,  but  act  simply  as  acids 
in  the  blood.  Acetone  does  not  have  this  effect,  not  being  an  acid, 
and  seems  not  to  be  toxic  to  any  considerable  degree ;  doses  of  4  grams 
per  kilo  cause  effects  similar  to  ethyl  alcohol  in  dogs,  8  grams  per 
kilo  being  fatal,  which  corresponds  to  a  dose  of  500  grams  for  an  adult 
man.  According  to  Rhann' ^"  acetone  is  more  toxic  (for  guinea  pigs) 
than  methyl  alcohol,  while  for  rabbits  Desgrez  and  Saggio  ^*  found 
acetone  the  least  toxic  of  the  acetone  bodies,  diacetic  acid  next,  and 
^-oxybutyric  acid  most.  Ehrmann  ^^  also  claims  that  he  has  pro- 
duced typical  coma  with  the  sodium  salts  of  butyric  and  of  /?-oxybuty- 
ric  acid,  but  as  high  as  40  grams  of  ^S-oxybutyric  acid  have  been 
found  in  the  day's  urine  of  a  non-diabetic  without  any  evidence  of 
intoxication.  Ewing  suggests  that  the  acetone  bodies  may  cause  renal 
injury,  which  is  usually  evident  in  acidosis,  and  M.  H.  Fischer's 
views  on  the  relation  of  acids  to  nephritis  accord  wdth  this  fact. 
The  withdrawal  of  the  inorganic  bases,  especially  Ca  and  Mg,  may 
also  be  responsible  for  symptoms,  as  it  is  well  established  that  a  proper 
balancing  of  these  ions  is  necessary  for  normal  cell  activity,  especially 
in  the  nervous  system. *° 

Hurtley  -^  sums  up  the  evidence  on  the  toxicity  of  the  acetone 
bodies  by  saying  that  aceto-acetic  acid  seems  to  be  highly  toxic  only 
in  depancreatized  animals,  while  oxybutyric  acid  is  practically  non- 
toxic. He  favors  the  view  that  aceto-acetic  acid  poisoning  is  respon- 
sible for  diabetic  coma,  for  it  increases  in  the  urine  on  the  approach 
of  coma,  and  the  ratio  of  aceto-acetic  to  butyric  acid  in  the  urine  in- 
creases with  the  severity  of  the  intoxication.  The  increased  propor- 
tion of  aceto-acetic  acid  presumably  means  that  it  is  being  produced 
in  such  quantities  throughout  the  body  that  the  liver  cannot  reduce 
as  large  a  proportion  to  oxj^butyric  acid  as  it  normally  does.  In  con- 
sidering the  possibility  that  the  acetone  bodies  may  be  responsible  for 
at  least  part  of  the  intoxication  of  diabetic  coma  we  must  bear  in 

36  Pribram  and  Loewy  (Zeit.  klin.  Med.,  101.3  (77).  .384)  siisifrost  that  ab- 
normal products  of  protein  cleavage  are  responsible,  and  Rosenblonm  (X.  Y.  ^led. 
Jour.,  Aug.  7,  1915)  reports  cases  of  typical  dialietic  coma  witlioiit  acetone  bodies 
in  the  urine. 

3 T  .Tour.  Amer.  :\red.  Assoc,  1012   (.58).  628. 

ssCompt.  Rend.  Soc.  Biol.,  1007    (63),  288. 

soBerl.  klin.  Woch.,  1013    (50),  11. 

40  See  Cammidge,  Amer.  Med.,  lOlG   (11),  363. 


554  AliM)RMAIATn:i<    IS    METABOLISM 

mind  that  the  evidence  of  their  low  toxicity  is  based  on  short  time 
experimental  intoxications,  and  that  they  may  be  found  to  be  much 
more  toxic  than  is  generally  assumed  when  they  are  allowed  to  act 
for  many  days  and  weeks  on  the  nervous  tissues,  as  they  do  in  dia- 
betes. That  is,  the  experimental  evidence  concerning  the  toxicity  of 
the  acetone  bodies  has  not  been  obtained  under  conditions  comparable 
to  those  of  diabetic  acidosis. 

Origin  of  the  Acetone  Bodies. — The  chemical  nature  of  the  acetone 
bodies  is  such  that  they  mijiht  readily  be  produced  from  any  or  all  of 
the  three  classes  of  foodstuffs. 

They  might  be  derived  from  carhohydrates,  as  is  the  closely  related 
lactic  acid,  but  we  know  that  this  is  not  the  usual  source.  On  the 
contrary,  administration  of  a  proper  amount  of  carbohydrates  under 
certain  conditions  may  cause  the  acids  to  disappear  from  the  urine, 
and  acetone  bodies  may  be  eliminated  in  large  quantities  while  the 
patient  is  on  a  diet  almost  free  from  carbohydrates.  Carbohydrates 
are,  indeed,  the  most  active  agents  in  preventing  the  formation  of 
these  ketone  bodies,  i.  e.,  they  are  antiketogenic.*' 

They  might  readily  be  formed  from  proteins  through  splitting  out 
of  the  NH2  group  from  the  amino-acids ;  indeed  the  amino-aeids 
are  generally  considered  as  a  source  of  the  acetone  bodies,*^  par- 
ticularly because,  whenever  there  is  considerable  pathological  break- 
ing-down of  proteins,  these  bodies,  especially  acetone,  may  appear  in 
the  urine ;  e.  g.,  during  absorption  of  exudates,  in  carcinoma,  and  in 
starvation  or  other  conditions  with  great  wasting  of  the  tissues. 
Dakin  ^*  has  shown  that  only  leucine,  histidine.  phenylalanine  and 
tyrosine  yield  diacetic  acid  when  perfused  through  the  liver,  while 
most  of  the  other  animo-acids  are  able  to  yield  sugar  in  diabetic  ani- 
mals, and  hence  are  antiketogenic. 

On  the  other  hand,  the  amount  of  acids  sometimes  found  in  the 
urine  seems  to  be  greater  than  can  be  explained  by  the  protein  de- 
struction that  occurs  (Magnus-Lev^O,'*^  and  in  diabetes  it  is  often  ob- 
served that  feeding  of  fats  and  fatty  acids  increases  the  output  of 
acetone  bodies,  and  hence  it  is  evident  that  acetone  bodies  may  be  de- 
rived from  the  fats.  /?-oxybut}Tic  acid  can  be  readily  produced  from 
fatty  acids,  especially,  of  course,  from  butyric  acid,  and  we  usually 
observe  an  increase  in  the  acetone  excretion  in  a  diabetic  given  large 
quantities  of  butter.  Other  higher  fatty  acids  are  also  found  to  cause 
increased  acetone  excretion. 

41  Cnnceriiiii<r  antiketofionosis  see  Woodvatt,  Jour.  Amor.  ]\ro(l.  Assoc,  1910 
(.55),  210!). 

■»■■!  Krnhdeii  and  liis  associates  liave  (Hofmeister's  Beitr.,  lOOfi  (8),  121:  lOOS 
(111,  11.  7- it  I  <li'iiioiistrat('d  that  the  liver  can  form  acetone  from  many  snli- 
slaiices  perfused  throujjh  it  in  tiie  blood,  includinfr  not  only  amino-acids  of  Die 
fatt\'  acid  scries,  hut  also  the  aromatic  radicals  of  the   jirotciii   molecule. 

4-«".T<>ur.  P.iol.  Cliem.,  lOlS   fl4),  .128. 

45  Arch.  c\p.  i'atli.  u.  I'harni..  1S90  (42),  149;  Erjrch.  inn.  Aled..  1908  (1). 
374. 


omais  OF  THE  acetone  bodies  555 

The  studies  of  Knoop,'"  and  his  associates  have  indicated  that 
ill  the  catabolisni  of  fatty  acids,  the  chains  are  bi'okeii  (hjwn  by 
oxidation  of  the  cafboii  atom  lliii'd  ffoiii  tlic  ciid,  tliat  is,  the 
^-position,  and  the  two  end  carbon  atoms  are  tlien  split  off.  Tliere- 
fore,  two  carbon  atoms  are  always  split  off  at  a  time,  and  hence  every 
fatty  acid  wliich  contains  an  even  numl)er  of  carl)ou  atoms  can  l^e 
oxidized  into  oxybntyric  acid,  which  includes  the  ordinary  fatty  acids 
(oleic,  palmitic  and  stearic)  of  fat  tissue,  which  have  each  an  even 
number  of  carbon  atoms  (16  or  18),  and  also  butyric,  caproie  and  sim- 
ilar acids.  Normal  fatty  acids  which  contain  an  odd  number  of  car- 
bon atoms  cannot  yield  oxybutyric  acid.  However,  according  to  A. 
Loeb,*'  aceto-acetic  acid  may  be  built  up  from  acetic  acid  in  the 
liver,  and  the  urine  in  diabetes  may  contain  acetic  acid.  "The  for- 
mation of  oxybutyric  acid  and  of  diaeetic  acid  in  all  these  cases  may 
be  said  to  be  due  to  the  fact  that  the  diabetic  oroanism  is  not  able 
quite  to  finish  the  attack  on  the  beta-carbon  atom  of  butyric  acid" 
(Folin). 

From  the  results  of  tliese  studies  it  seems  that  the  acetone  bodies 
can,  theoretically  be  formed  from  any  of  the  three  classes  of  food- 
stuffs, but  that  ordinarily  they  come  chiefly  from  the  fats,  and  in 
severe  diabetes  also  to  considerable  extent  from  fatty  acids  formed 
hy  deaminization  of  amino-acids.  Although  it  is  probable  that  the 
acetone  bodies  are  formed  in  many  if  not  all  tissues,  yet  there  is 
abundant  evidence  that  the  liver  plays  an  important  part  in  ketogene- 
sis,  as  shown  by  the  decrease  in  acetone  bodies  in  Eck  fistula  dogs,  and 
their  great  increase  when  the  blood  supply  of  the  liver  is  augmented.*^ 

In  addition  to  the  sources  of  acidosis  substances  from  metabolic  proc- 
esses, as  outlined  above,  it  is  also  possible  that  they  may  be  derived 
from  organic  acids  formed  by  bacterial  action  in  the  alimentary  canal, 
as  emphasized  by  Palacios.^*'' 

Sarcolactic  Acid  often  is  found  in  the  urine,  but  in  origin  and  sig- 
nificance it  is  entirely  different  from  the  acetone  bodies,  and  it  prob- 
ably is  never  present  in  sufficient  amounts  to  cause  an  acid  intoxica- 
tion by  abstraction  of  alkalies  from  the  blood.  //(  vitro,  we  obtain 
sarcolactic  acid  whenever  sugar  is  placed  in  an  alkaline  solution, 
provided  the  supply  of  oxygen  to  the  solution  is  deficient:  but  if  the 
oxygen  supply  is  adequate,  sugar  will  not  yield  lactic  acid  with  alka- 
lies (Nef).  Similarl}',  an  isolated  surviving  muscle,  when  asphyxi- 
ated by  any  means,  shows  a  rapid  accumulation  of  lactic  acid,  which 
fails  to  occur  when  sufficient  oxygen  is  supplied.  This  lactic  acid 
comes  chiefly  from  sugar,  but  about  25  to  ;30  per  cent,  of  it  can  have 
it:;  origin  in  protein    (or  fat?)    (Woodyatt).     If  an   organism   as  a 

40  Full  bibliofirraphy  and  discussion  by  Porges.  Frsjcbnisse  Physiol..   1010    (10), 
G.     See  also  Rinfrer,  Jour.  Piol.  (hem.,  1!)1.3,  Vol.  14. 
4V  Biochem.  Zeit.,   1012    (47),   US. 

48  Fischer  and  Kossow.  Dent.  Arch.  klin.  INfed..  101.3    (101).  470. 
48aAmer.  Jour.  Med.  Sci.,  1015   (149),  267;  Med.  Eecord.  starch  25,  1916. 


556  ABXOliMALITIES    IX    METABOLIHU 

whole  is  iiisufifieiently  supplied  with  oxygeu,  lactic  acid  accumulates 
in  the  tissues  and  appears  in  the  urine,  disappearing-  when  the  oxy- 
gen supply  is  restored.  Lactic  acid  often  appears  after  poisoning 
with  a  large  number  of  drugs,  which  Loewy  has  classified  as  drugs 
whose  action  in  the  hody  resembles  that  of  lack  of  oxygen  (arsenic, 
phosphorus,  hydrazine,  chloroform,  etc.).  These  poisons  are  all  char- 
acterized by  causing"  impoverishment  of  glycogen,  fatty  liver,  and 
acute  degenerative  changes  especially  in  the  liver  cells  and  the  endo- 
thelium. Therefore  the  assumption  seems  justified  that  the  poisons 
and  conditions  which  lead  to  lactic  acid  excretion  depend  ultimately 
upon  impairment  of  the  interchange  of  oxygen  in  the  cells.  Wood- 
yatt  states  that,  so  far  as  known,  lactic  acid  has  never  been  demon- 
strated in  any  tissue  in  which  deficient  oxygenation  can  be  excluded, 
and  regards  lactic  acid  as  the  metabolite  of  asphyxia  or  its  equivalent. 
Over  against  this  view  is  that  of  Embden  and  his  associates,  which 
is  shared  b^-  others,  that  lactic  acid  is  a  normal  intermediary  in  the 
breakdoA\Ti  of  the  sugars  in  the  body,  its  direct  antecedent  being  a 
triose,  but  perusal  of  their  work  only  emphasizes  that  in  all  the  con- 
ditions in  which  their  data  were  obtained  asphyxial  conditions  were 
present;  furthermore,  this  conception  of  lactic  acid  as  a  chief  inter- 
mediate in  normal  sugar  catabolism  is  not  in  harmony  with  the  best 
ideas  of  carbohydrate  chemistrj^  (Woodyatt).  This  author  has  fur- 
thermore found,  by  direct  observation  of  the  utilization  of  lactic  acid 
wdien  injected  intravenously,  that  it  cannot  well  be  an  important  inter- 
mediate in  carbohydrate  catabolism. ^^"^ 

It  is  possible  that  the  presence  of  lactic  acid  in  the  urine  may  also 
result  from  defective  transformation  of  ammonia  into  urea  by  a  dis- 
eased liver,  the  acid  neutralizing,  and  being  excreted  with,  the  am- 
monia; in  this  case  no  defective  oxidation  need  be  assumed.  How- 
ever, administration  of  phlorhizin  to  phosphorus  poisoned  dogs  causes 
both  ammonia  and  lactic  acid  to  disappear  from  the  urine,  indicating 
that  the  ammonia  is  the  protective  substance  which  neutralizes  the 
lactic  acid,  and  not  the  reverse. 

Sarcolactic  acid,  which  is  dextrorotary,  must  be  distinguished  from 
its  optical  isomer,  the  inactive  lactic  acid  that  is  produced  by  fer- 
mentation. AVhen  this  fermentation  lactic  acid  is  formed  in  the  stom- 
ach and  enters  the  blood,  it  ordinarily,  like  other  ingested  organic 
acids,  is  combined  by  the  blood  alkalies  and  oxidized  to  carbonates. 
It  is  doubtful  if  it  ever  enters  the  urine.*" 

As  a  general  rule  sarcolactic  acid  is  not  found  abundant  in  the 
urine  together  with  the  acetone  bodies,  but  is,  indeed,  antiketogenic. 
Its  appearance  in  the  ui-ine  indicates  tliat  glycogen  is  not  com]ilctoly 

■•^th  TTarvey  Society  Lectures,   lOlG. 

<"  Tlie  theory  of  Boix  that  cirrhosis  of  (lie  liver  may  he  jirodiiced  by  butyric 
acid  formed  in  fiastric  ferment^Ttictn  coiild  im<  lie  corroborated  bv  Joaniiovics, 
Arch.  int.  Pharmacodyn.,   190.')    (l.'j),  241. 


ACID  IXTOMCAT/OX  I\  XOXDIAnHTKJ  COXDITIOyH  557 

burned,  and  this  condition  is  usually  aeconipanied  witJL  fatty  changes 
in  the  liver,  which  also  depend  on  lack  of  oxidation.  Throughout  the 
clinical  forms  of  acidosis,  lactic  acid  and  fatty  degeneration  are 
always  associated  (Ewiug).  To  assume,  as  has  been  generally  done, 
that  the  lactic  acid  appears  in  the  urine  when  hepatic  alterations  are 
marked,  because  of  a  loss  of  the  liver  tissue  which  should  destroy  it, 
is  probably  not  warranted.  Rather,  the  liver  conditions  and  the  for- 
mation of  lactic  acid  depend  upon  the  same  cause,  which  is  a  de- 
fective oxygen  supply  or  interchange,  either  general  or  local. ''''"^ 

ACID  INTOXICATION  IN  CONDITIONS  OTHER  THAN  DIABETES  «b 

Clinical  Types  of  Acidosis. — Ewing  has  divided  acidosis  as  it  oc- 
curs in  man,  into  two  main  types:  1.  Acidosis  resembling  in  its 
effects  the  acidosis  produced  by  experimental  injection  of  acids. 
This  is  characterized  by  the  excretion  of  the  acetone  bodies  in  large 
amounts  with  a  corresponding  amount  of  ammonia  in  the  urine, 
and  by  the  absence  of  marked  and  characteristic  anatomical  changes. 
Diabetic  acidosis  and  the  acidosis  of  starvation  are  the  clinical  condi- 
tions showing  this  type. 

2.  Acidosis  resembling  that  produced  experimentally  by  extirpa- 
tion of  the  liver  or  by  the  Eck  fistula.  Here  the  urine  contains  much 
lactic  acid  and  relatively  little  acetone  bodies,  the  ammonia  being 
in  excess  of  any  acetone  compounds,  and  there  is  also  much  incom- 
pletely changed  nitrogenous  compounds.  The  clinical  prototj^pes  are 
phosphorus  and  chloroform  poisoning,  the  toxemias  of  pregnancy  and 
cyclic  vomiting  and  acute  yellow  atrophy.  Anatomically  it  is  char- 
acterized by  severe  hepatic  degeneration,  usually  fatty. 

This  classification  is  purely  one  of  convenience,  for  typical  and  ex- 
treme fatty  liver  may  occur  in  phlorhizin  diabetes  with  the  typical 
coma,  provided  the  animal  was  fat  when  the  experiment  was  begun, 
and  that  the  experiment  was  not  carried  on  in  such  a  way  as  to  ex- 
haust the  fat  temporarily  present  in  the  liver.  It  is  tnie  that  high- 
grade  fatty  livers  are  not  usually  observed  in  most  cases  of  long- 
standing diabetes,  but  fatty  changes  are  severe  in  the  acute  fulmi- 
nating types  of  diabetes  of  infancy  and  childhood,  and  here  we  have 
typical  acidosis.  The  reason  that  lactic  acid  does  not  occur  in  the 
urine  of  a  completely  diabetic  animal  or  man,  is  probably  not  be- 
cause of  any  assumed  destructive  power  of  the  comparatively  nor- 
mal liver,  but  because  here  the  carbohydrate  equilibrium  is  so  dis- 
turbed that  even  if  lactic  acid  were  formed  it  would  necessarily  be 
converted  into  glucose  and  appear  in  the  urine  as  such.  ]\Iandel 
and  Lusk  have  indeed  shown  that  even  dogs  poisoned  with  phos- 
phorus excrete  no  lactic  acid  in  the  urine  if,  in  addition,  they  are 

49a  See  Macleod  and  Wedd  (.Tour.  Biol.  Clicm.,  1014  (IS),  446)  wlio  found 
that  reducing  the  oxygen  supply  to  the  liver  caused  a  marked  rise  in  the  lactic 
acid  content  of  the  hepatic  blood. 

49b  See  resuna^  by  Frothingham,  Arch.  Int.  Med.,  1916   (IS),  717. 


558  AIi\ORMALITli:s    I\    METAliOIAHM 

made  completely  diabetic  Avith  phlorliizin,  and  that  lactic  acid  in- 
jected subcutaneoiisly  into  a  phlorliizinized  animal  is  excreted  gram 
for  o-ram  as  sugar. 

Not  infrequently  acetone  and  diacetic  acid,  less  often  oxybutyric 
acid,  are  found  in  the  urine  of  patients  suffering  from  the  most  di- 
verse diseases.  It  is  customary  to  refer  to  this  condition  as  " aceto- 
tieniia"  or  "acetomiria,"  and  to  ascribe  many  of  the  observed  symp- 
toms to  "acid  intoxication."  The  presence  of  these  substances  in  the 
urine,  however,  is  by  no  means  evidence  of  acidosis,  for  excretion  of 
considerable  amounts  of  acetone  bodies  may  occur  Avithout  reduced 
COg-carrying  capacity'  of  the  blood,  and  they  may  be  absent  with 
marked  acidosis.  In  addition,  it  must  be  kept  in  mind  that  acidosis 
may  result  from  other  causes  than  over-production  of  organic  acids; 
e.  g.,  acid  phosphate  retention  in  nephritis,  or  loss  of  bases  from  biliary 
or  pancreatic  fistula.  In  no  other  condition  do  the  amounts  of  organic 
acids  in  the  urine  approximate  the  amounts  found  in  diabetic  coma. 
Therefore,  the  intoxication  in  these  cases  is  probably  not  due  to  the 
acids,  but,  on  the  contrary,  the  presence  of  the  acetone  bodies  is  due 
more  often  to  the  effects  of  toxic  substances  of  diverse  origins  and  na- 
tures. 

Anesthesia. — As  shown  first  by  Greven  (1895),  and  especially  by 
Brewer  and  by  Helen  Baldwin,^"  acetone  is  nearly  always  present  in 
the  urine  during  the  first  twenty-four  hours  after  administration  of 
either  chloroform  or  ether,  and  occasionally  diacetic  acid  appears  on 
the  second  or  third  day  after;  but  ordinarily  there  is  no  increase  in 
organic  acids  in  the  urine.  The  starvation  preceding  and  following 
the  operation  is  also  a  factor  of  considerable  importance.  It  does  not 
seem  probable  that  the  symptoms  observed  in  typical  cases  of  delayed 
chloroform-poisoning  are  due  chiefly,  if  at  all,  to  acid  intoxication 
per  se,  but  rather  are  the  result  of  extensive  injury  to  the  parenchy- 
matous organs,  particularly  the  liver,  by  the  chloroform,  which  causes 
a  condition  resembling  acute  yellow  atrophy  or  phosphorus-poison- 
ing.^^ 

Cachectic  Acetonuria. — Acetone  and  diacetic  acid,  but  less  abun- 
dantly the  oxybutyric  acid,  are  found  in  the  urine  in  many  condi- 
tions associated  with  wasting,  among  Avhich  may  be  especially  men- 
tioned : 

Infantile  marasmus,"-  in  which  increased  ammonia  is  found  in  the 
urine,  and  sometimes  sym])toms  resembling  acid  intoxications  occur. 
Normally  the  urine  of  suckling  infants  contains  1-4  mg.  per  day  of 
acetone  bodies,  which  may  be  increased  to  15-35  mg.  by  simple  hun- 
ger. In  fact,  "acidosis"  seems  to  occur  particularly  frequently  in 
infants   from   relatively   slight   causes,   such    as   gastro-enteritis   and 

■in  .I.Hir.  .,f  T^iol.  Choni.,  lOOG    (1),  239. 

•"'i  Wells,   .lour.   Amcr.   ]\lcd.    Assoc.    l!)(l)i    ()('.].    :M1. 

52  See  Meyer  and  Laii{i;stein.  .Jalirb.  f    Isiiidrrhcilk..  1!M)(1    (fi.T).  .10. 


\()\-I)/\I!i:tic  Ac/n  nrox/r  \'ni)\  559 

other  infectious  conditions.  'J'liis  is  perliaps  due  to  a  lower  oxidizing 
power  on  the  part  of  the  infantile  organism  ( Pfaundler),''^  since 
the  proportion  of  nitrogen  in  the  urine  of  infants  in  forms  other 
than  urea,  is  higher  than  in  adults.  Even  an  unusually  fatty  diet 
may  cause  aeetonuria  in  infants.  It  has  also  been  suggested  that  ex- 
cessive formation  of  acids  in  the  intestine  through  bactei-ial  decompo- 
sition may  cause  withdrawal  of  tlic  bases  from  tlic  IjIooiI.  wliich  are 
lost  to  tlie  body  through  excretion  in  the  feces. 

Starvation. — Acetone,  which  is  normally  excreted  through  the 
lungs  for  tlie  most  part  (80-90  per  cent,  of  that  produced)  appears 
in  excess  in  the  urine  verj'  soon  after  fasting  begins,  there  being 
more  produced  than  can  be  exhaled.  After  24  to  36  hours  of  fasting, 
diacetic  acid  appears,  and  then  oxybutyric  acid,  which  may  reach  10 
to  20  grams  per  day  in  starvation,  and  even  higher  figures  are  re- 
corded. The  urinary  ammonia  nitrogen  runs  parallel  to  the  acidosis. 
The  use  of  a  carbohydrate-free  diet  is  also  accompanied  by  a  marked 
acetonuria,^^''  no  matter  how  much  fat  is  supplied,  which  may  reach  a 
point  where  several  grams  of  oxybutyric  acid  are  being  excreted  per 
day  without  symptoms  of  serious  intoxication.  A  relatively  small 
amount  of  carbohydrate  (80  grams)  is  sufficient  to  prevent  this 
acidosis.  If  the  meat-fat  diet  is  continued  for  some  time,  however, 
there  seems  to  be  some  sort  of  adaptation  so  that  the  aeetonuria  di- 
minishes until  practically  normal  figures  may  be  reached. 

Acidosis  of  Pre^ancy. — During  pregnancy  the  urine  usually  con- 
tains acetone  in  slight  excess,  and  occasionally  is  in  large  excess  in 
women  who  are  suffering  from  the  toxemias  of  pregnancy.  Here 
there  is  a  rise  in  ammonia  far  beyond  the  proportion  of  acetone  bod- 
ies, partly  because  of  the  large  amounfs  of  lactic  acid  which  are  ex- 
creted, and  partly  from  abnormal  protein  metabolism  and  tissue  de- 
struction, but  the  proportion  of  the  urinarv^  nitrogen  which  is  consti- 
tuted by  ammonia  is  too  inconstant  to  serve  as  a  prognostic  and 
operative  guide.  Ewing  has  observed  a  case  of  pernicious  vomiting 
with  75  per  cent,  of  the  total  nitrogen  as  ammonia,  and  no  urea, — 
while  there  may  occur  fatal  cases  without  large  excess  of  ammonia. 
Higher  ammonia  figures  are  usually  reached  in  pernicious  vomiting  of 
pregnancy  than  in  eclampsia ;  in  neither  is  the  acidosis  present  suffi- 
cient to  account  for  the  intoxication.  (See  discussion  of  "Eclamp- 
sia.") Even  normal  pregnant  women  seem  to  show  a  reduced  ability 
to  tolerate  a  deficiency  in  the  carbohydrates  of  the  diet.^* 

Cyclic  Vomiting. — Here  the  urine  usually  shows  acetone  bodies,  lac- 
tic acid,  indican  in  excess,  and  a  rise  in  the  proportion  of  neutral 
to  oxidized  sulphur  (Howland  and  Richards).  As  these  findings 
may  persist  in  spite  of  absorption  of  carbohydrates,  they  are  not  en- 

33,Jahrb.  f.   Kinderheilk..   1001    (.54),   247. 

53a  See  Higgins.  Peabodv  and  Fit/.  Jour.  ^Fed.  T!os..  1910    (.341,  203. 

54Porges  and  Novak,  Berl.  klin.  Woch.,  1911    (4S),  I7")7. 


560  ABNORMALITIES    IX    METABOLISM 

tirely  due  to  starvation,  and  there  are  severe  fatty  changes  in  the 
liver  and  kidneys,  indicating  a  toxemic  origin  associated  with  de- 
fective oxidation.  ^Iclhinl)y  ■'"'  fuuntl  a  considerable  creatine  elimina- 
tion in  a  typical  case,  together  with  the  acidosis. 

Inanition  and  Cachexia^ — Under  this  heading  may  be  grouped  the 
acetonuria  observed  in  intestinal  disturbances  in  children,'^^'^  hys- 
terical vomiting,  psychoses,  and  cancer.  In  each  of  these  conditions 
coma  of  the  type  of  diabetic  coma  has  sometimes  been  observed,  and 
in  all  of  them  acetonuria  is  common,  the  reasons  being  obvious  after 
the  above  discussion.  A  relative  acidosis  may  also  result  from  de- 
ficiency of  bases  in  the  diet  of  growing  infants.  In  many  cases  of 
acidosis  of  infants  there  is  not  sufficient  increase  in  the  acetone  bodies 
of  the  blood  to  account  for  the  acidosis ;  "'^  on  the  other  hand,  most  of 
the  children  excreting  acetone  bodies  in  the  urine  do  not  have  acidosis. 

Retention  of  placenta  or  fetus,  acetonuria  being  considered  of  di- 
agnostic value  in  determining  the  death  of  the  fetus  in  utero,^''  but 
not  in  extrauterine  pregnancy  (Wechsberg).-'* 

In  uremia,  as  previously  mentioned,  organic  acids  may  appear  in 
the  urine,  but  apparently  as  a  result,  and  not  as  the  cause,  of  the 
uremia  (Orlowski).  There  is  usually  some  acidosis  in  advanced 
nephritis,  but  marked  only  in  uremia. ^*^  Here,  according  to  ]\Iarriott 
and  Howland,^^''  acid  phosphates  which  the  kidney  has  failed  to  ex- 
crete, may  be  an  important  factor.  Sometimes  in  advanced  nephritis 
the  acidosis  may  be  of  such  a  degree  as  to  simulate  diabetic  coma,  and 
the  nocturnal  hyperopnoea  of  nephritis  probablj"  is  the  result  of 
acidosis  (Whitney-^). 

Other  Conditions. — Acetonuria  is  observed  inconstantly  in  fever, 
especially  in  children;  also  after  poisoning  by  many  drugs,  includ- 
ing, besides  the  heavy  metals,  morphine,  atropine,  antipyrine,  and 
phlorhizin.  Asiatic  cholera  shows  a  marked  acidosis,  in  manj^  re- 
spects resembling  diabetic  acidosis,'**'^  and  gastro-intestinal  infections 
of  similar  sorts  may  cause  severe  acetonuria.  "Whitney  -^  finds  acid- 
osis to  be  a  common  terminal  event  in  many  diseases,  and  often  the 
immediate  cause  of  death.  Pneumonia  is  accompanied  by  acidosis,^*^ 
often  of  serious  degree,  subsiding  rapidly  after  the  crisis.  At  high 
altitudes  there  is  always  an  acidosis,  which  stimulates  the  respiratory 
center  to   increased   activity.     In   asphyxial   conditions   of    all   sorts 

S5  Lant'ot,  Jiilv   1,   UMl. 

55a  See  Ilowland  and  .Marriott,  Amer.  Jour.  Dis.  Child..  lOlC,  (11),  ,?0!) :  (12), 
459. 

■">'!  Moore,  Amcr.  .Tour.  Dis.  Cliild.,  miC,    (12),  244. 

57  See  Frojumer.  Borl.  Ulin.  Woch.,  100;!    (42),  1008. 

ssWien.  klin.  Wocli.,  1000    (10),  053. 

r.'^aSee  Scllards,  Hull.  Jolins  Hopkins  Hosp.,  1014  (2.")).  141:  Toabodv,  Arch. 
Int.  :Med.,    1015    (10),  455. 

58i>  Arcli.  Int.  :M('d.,  lOlG  (IS),  70S. 

f'Sc  Sellards  and  Sluikloo.  Pliilipi)ine  Jour.  Sci.,  1011    (G),  53. 

50  Lewis  and  Barcroft,  Quart.  Jour.  Med..   l!)15    (S),   lOS. 


FATiaUE  5G1 

aeiduisis  is  present,  c.  g.,  uncompensated  cardiac  defects,  severe  anemia, 
gas  poisoning. 

FATIGUE 

The  symptoms  of  fatigue,  whether  general  or  local,  seem  to  be  due 
to  an  intoxication  with  the  products  of  the  excessive  metabolic  activ- 
ity, and  part  of  the  symptoms,  at  least,  seem  to  be  due  to  acid  intox- 
ication. Among  the  metabolic  products  of  muscular  activity  are 
known  to  be  creatin,  creatinin,  sarcolactic  acid,  and  carbon  dioxide. 
The  amount  of  acid  developed  in  an  active  muscle  is  quite  consider- 
able, and  when  the  activity  is  violent  or  prolonged  the  sarcolactic 
acid  accumulates,  being  formed  faster  than  it  can  be  removed.  Part 
of  the  acidity  of  the  muscle  is  due,  however,  not  to  the  sarcolactic 
acid  itself,  but  to  monopotassium  phosphate  (KHoPO^),  which  is 
formed  by  the  action  of  the  sarcolactic  acid  upon  the  dipotassium 
phosphate  present  in  the  blood  and  muscle.  The  effect  of  these 
various  substances  upon  muscular  fatigue  has  been  studied  experi- 
mentally, and  while  the  creatin  seems  not  to  be  a  "  fatigue  substance, ' ' 
sarcolactic  acid,  monopotassium  phosphate,  potassium  sarcolactate, 
and  carbon  dioxide  all  cause  muscle  tissue  to  react  to  stimuli  in  the 
same  way  that  a  fatigued  muscle  does  (Lee).*"^ 

It  is  quite  probable  that  the  muscular  weakness  of  diabetics,  and 
the  exhaustion  associated  with  many  conditions  in  which  organic 
acids  appear  in  the  urine  in  abnormal  quantities,  depend,  at  least  in 
part,  upon  the  effect  of  these  acids  upon  the  muscle  tissue,  for  Lee 
found  that  |8-oxybutyric  acid  causes  the  same  fatigue  reaction  in  mus- 
cles as  does  sarcolactic  acid.  Furthermore,  sarcolactic  acid  itself 
often  appears  in  the  urine  in  these  conditions.  It  may  be  added  that 
in  fatigued  animals  the  alkalinity  of  the  blood  (by  titration)  has  been 
found  decreased  (Geppert  and  Zuntz),  and  the  proportion  of  the 
urinary  nitrogen  that  appears  in  other  combinations  than  urea  is 
increased  (Poehl).'^^ 

The  "Toxins"  of  Fatigue. — In  extreme  exhaustion  the  evidences 
of  a  general  intoxication  often  become  severe,  so  that  the  condition 
may  resemble  an  acute  febrile  disease  and  last  for  several  days.  It 
seems  very  probable  that  substances  more  toxic  than  the  above-men- 
tioned acids  are  involved.  Weichardt '-  claims  to  have  demonstrated 
as  produced  by  muscular  fatigue  a  toxic  substance,  which  in  structure 
resembles  the  bacterial  toxins,  called  by  him  kenotoxin,'^-"'  and  against 
which  an  antitoxin  may  be  obtained.  This  toxic  material  is,  he  be- 
lieves, formed  from  the  protein  molecule  in  the  first  stages  of  its  de- 

60  Jour.  Amer.  Med.  Assoc,  1906  (46),  1401;  where  is  given  a  complete  review 
of  the  subject  of  fatigue,  with  tiie  literature. 

eiDeut.  med.  Woch.,   1001    (27),  706. 

«2  "Ueber  Enniidungsstoffe,"  Enke,  Stuttgart,  1012;  Kolle  and  Wassermann's 
Handbuch,    1013    (2),    1400. 

02a  See  Weichardt  and  Schwenk,  Zeit.  phvsiol.  Chem..  191.3   (83),  381. 
36 


562  ABXOKMALITIES    IX    METABOLISM 

composition,  as  a  side  product  which  is  normally  protected  against 
by  a  formation  of  an  antitoxin,  rather  than  bj^  being  split  up  further^ 
as  is  the  case  with  the  rest  of  the  protein  molecule.  It  is  excreted  not 
only  in  the  urine,  but  also  in  the  breath,  and  may  be  produced  in  vitro 
by  disintegrating  protein  at  temperatures  under  40°.  Various  poisons, 
cause  kenotoxin  to  appear  in  the  urine,  and  it  is  found  in  the  urine 
of  many  animals,  as  well  as  in  plant  tissues.**^  The  study  of  anaphy- 
laxis has  led  to  so  many  evidences  of  the  remarkable  toxicity  of  the 
products  of  protein  cleavage,  that  the  possibility  that  some  of  these 
may  be  responsible  for  fatigue,  as  Weichardt  has  so  vigorously  main- 
tained, is  receiving  much  support."*  Whether  this  work  is  coiitirmed 
or  not,  there  remains  the  fact  that  the  blood  of  fatigued  animals 
contains  toxic  substances,  which  ^losso  proved  as  follows:  Tf  blood 
is  transfused  from  an  exhausted  dog  to  a  normal  dog,  from  which 
an  equivalent  amount  of  blood  has  been  withdrawn,  this  second  dog^ 
will  show  the  usual  manifestations  of  fatigue. 

Mental  Fatigue. — The  chemical  changes  of  mental  fatigue  are  not 
known,  but  it  is  known  that  the  ganglion-cells  show  marked  struc- 
tural alterations  as  a  result  of  fatigue,  chromatolysis  often  being 
very  striking.  Since  lecithin  forms  so  important  a  part  of  the  nerv- 
ous system,  it  is  tempting  to  imagine  that  in  fatigue  excessive  quan- 
tities of  its  toxic  decomposition-product,  choline,  and  the  still  more 
toxic  derivative  of  choline,  neurine,  are  formed  in  considerable 
amounts  and  cause  part,  at  least,  of  the  intoxication. 

That  choline  or  neurine  actually  are  the  cause  of  any  of  the  symp- 
toms of  fatigue,  however,  has  not  been  established ;  but  Donath  '^^ 
considers  choline  an  important  factor  in  the  production  of  epileptic 
convulsions J^*^  Animals  kept  for  a  long  time  from  sleeping  show  the 
presence  in  their  blood,  cerebro-spinal  fluid  and  brain  tissues,  of  a 
poisonous  property  causing  somnolence  in  other  animals  (Legendre 
and  Pieron).**'  This  cannot  well  be  choline  or  any  similar  substance, 
for  it  does  not  filter,  is  insoluble  in  alcohol,  and  is  destroyed  by 
heating  at  65°. 

THE  POISONS  PRODUCED  IN  SUPERFICIAL  BURNS  «* 

In  a  certain  proportion  of  cases  of  extensive  but  superficial  burns,, 
death  follows  after  an  interval  of  from  six  hours  to  a  few-  days,  ap- 

03  Cent.  f.  Bakt..  in07   (4.3),  .312. 

•■'♦  The  failure  of   various   investigators  to  corroborate  Woiiliardt    is  disciissod 
l)v  Konricli.  Zcit.  f.  TTvp.,  1014   (7S).  1:  Korflf-Petersen.  ihUJ..  p.  37. 
■  o^' Zeit.   pliysiol.  Chem..   1003    (.''0),   -,20. 

•"■•  Confeiiiin<r  tlie  theories  i 'vl  I'crature  of  tlie  subjeet  of  epilepsy  in  relation 
to  its  patliolofjical  eliemistry  aiu'  to  aiitoiiitoxieatioii.  s(H>  the  review  of  ^lasoin. 
Aroh.  internat.  de  Pharniaeodvnainie,  1004    (13),  3S7. 

«7  Zeit.  alljr.  Physiol.,   1012'  (14).  235. 

«s  Literature  piven  li\-  P.ardeen.  .Tolins  Hopkins  llosp.  Peports.  ISOS  (7).  137; 
Eyff,   Cent.    Orenzf^'el).   :\lc(l.    u.    Cliir..    1001     i4),    42S:    PfeillVr,   Virdiow's   Areh... 


I'uisoxs  I'Roui  ri:u  IS  jiLJtxs  563 

parently  because  of  a  profound  intoxication.  As  evidence  of  intoxica- 
tion we  Imve  not  oidy  clinical  manifestations,  such  as  delirium,  iiemo- 
^lobinuria,  and  albuminuria,  vomitinj>^,  bloody  diarrhea,  etc.,  but,  more 
convincinoly,  the  anatomical  lindings  at  autopsy,  which  are  strikingly 
similar  to  those  resulting  from  acute  intoxication  with  bacterial  prod- 
ucts. Uardeen  found  (juite  constantly  cloiuly  swelling  and  focal  and 
parenchymatous  degeneration  in  the  liver  and  kidneys:  softening  and 
enlargement  of  the  spleen  with  focal  degeneration  in  the  ^Iali)ighian 
bodies ;  and  particularly  degenerative  changes  in  the  lymph-glands 
and  intestinal  follicles  resembling  those  observed  in  diphtheria,  which 
^McCrae '••'  considers  due  to  proliferation  and  phagocytosis  by  the 
endothelial  cells  of  the  lymphatic  structures.  IMarked  changes  are 
usually  present  in  the  blood,  consisting  of  fragmentation  and  dis- 
tortion of  the  red  corpuscles,  hemoglobinemia,  loss  of  water  with  a 
relative  increase  in  the  number  of  corpuscles  by  from  one  to  four 
millions  per  cubic  millimeter,  an  increase  in  the  blood  platelets,  and  a 
rise  in  the  number  of  leucocytes  as  high  as  30,000  to  50,000.'^°  Hem- 
oglobinuria is  also  frequently  present,  and  almost  constantly  gastro- 
intestinal irritation  occurs,  with  anatomical  evidences  of  acute  enter- 
itis, acute  gastritis,  and  occasionally  gastric  or  duodenal  ulcers.  Ac- 
cording to  Korolenko,"^  the  sympathetic  nervous  system  is  seriously 
involved. 

It  therefore  seems  probable  that  poisons  are  formed  as  a  result 
of  superficial  burns,  which  have  the  effect  of  causing  hemolysis,  and 
which  are  also  cytotoxic  for  parenchymatous  cells  and  particularly 
for  nervous  tissues.  These  hypothetical  poisons  seem  to  be  eliminated 
by  the  intestines  and  kidneys,  which  are  injured  by  the  poisons  in 
their  passage  through  these  organs.  The  attempts  to  explain  all  the 
observed  effects  of  burns  as  due  to  thrombosis  or  to  embolism  by  al- 
tered red  corpuscles  seem  to  have  failed,  for  the  peculiar  location  of 
the  lesions  (e.  g.,  duodenal  ulcers,  necrosis  in  the  Malpighian  bodies 
of  the  spleen,  etc.)  does  not  agree  with  this  hypothesis,  and  there  are 
too  many  evidences  of  the  presence  of  some  decidedly  toxic  substance 
in  the  blood.  There  can  be  no  question  that  the  poisonous  substance 
or  substances  are  formed  in  the  burned  area,  and  not  in  the  internal 
organs  as  a  result  of  hyperpyrexia,  as  shown  by  numerous  observa- 
tions. Thus,  if  the  burned  area  is  removed  immediately  (in  narco- 
tized experimental  animals),  death  will  be  prevented,  whereas  if  the 
burned  tissue  is  permitted  to  remain  for  a  few  hours,  death  will 
occur.  If  the  burned  skin  is  transplanted  to  a  normal  animal,  this 
animal  will  develop  symptoms  of  intoxication,  while  the  burned  ani- 
mal may  be  saved  by  the  transplantation   (Vogt).     The  poison  ap- 

190.5   (180).  367.     Full  discussion  of  theories  by  Vopt,  Zeit.  exp.  Path.  u.  Pliarm., 
1912   (11),  191. 

69Amer.  :Med..  1901    (2),  735. 

TO  Locke,  Boston  :\led.  and  Surg.  Jour.,  1902  (147),  480. 

71  Cent.  f.  Path.,  1903  (10),  663. 


564  ABNORMALITIES    IX    METABOLISM 

pears  to  be  absorbed  from  the  burned  area  into  the  blood,  for  if  the 
circulation  is  shut  off  from  the  burned  area,  no  intoxication  results; 
this  probably  explains  in  part  why  deep  destructive  bums  of  small 
areas,  which  are  associated  with  local  thrombosis,  are  much  less  seri- 
ous than  a  superficial  slight  scalding  over  a  large  area.  Apparently 
the  poison  is  produced  chiefly  or  solely  in  the  skin,  for  burning  of 
muscle  is  not  followed  by  intoxication  (Eijkman  and  Hoogenhuyze).'- 
AVhen  one  of  a  pair  of  animals  united  to  another  by  operative  pro- 
cedure (parabiosis)  is  burned,  the  other  animal  may  become  intoxi- 
cated, while  the  intoxication  of  the  burned  animal  is  less  than  it 
would  be  if  it  were  alone  (Vogt). 

Numerous  investigators  have  reported  finding  poisonous  sub- 
stances in  the  blood,  tissues,  or  urine  of  burned  men  and  animals,  but 
the  reports  disagree  widely  in  details.'^  Thus  Dietrichs  states  that 
the  blood  of  burned  animals  contains  hemolysins  and  hemao:glutinins, 
which  could  not  be  corroborated  by  Burkhardt  ^*  or  b}^  Pfeiffer/' 
The  latter,  however,  finds  that  the  urine,  serum,  and  organs  of  burned 
animals  contain  substances  poisonous  for  the  same  and  for  dififerent 
species,  which  is  in  accord  with  the  results  of  numerous  earlier  inves- 
tigators. The  poisons,  according  to  Pfeiffer,  are  neurotoxic  and  necro- 
genic  in  their  properties,  and  act  without  a  period  of  incubation ; 
they  are  rapidly  w^eakened  on  standing  in  solution  and  by  the  action 
of  sunlight,  are  absorbed  from  the  gastro-intestinal  tract,  are  soluble 
in  water,  alcohol,  and  glycerol,  but  not  in  chloroform  or  ether,  are 
precipitated  by  HgCL  in  acid  solution,  and  by  phosphotungstic  acid, 
and  they  are  not  volatile.  Apparently,  according  to  Pfeiffer.  they 
are  not  ptomains,  nor  yet  pyridine  derivatives,  as  many  investigators 
have  contended,  but  resemble  more  closely  the  labile  poisons  of  snake 
venom,  and  have  effects  similar  to  the  unknown  poisons  that  are  con- 
cerned in  uremia.  The  neurotoxic  substance  is  more  thermostable 
than  the  necrogenic  substance,  which  is  very  easily  destroyed  by  heat. 
Pfeiffer  believes  it  probable  that  the  poisons  are  derived  from  the 
cleavage  of  proteins  altered  in  composition  by  burning,  and  he  finds 
an  enzyme  splitting  glj'cyltryptophane  in  the  blood  and  urine  of 
burned  animals."'"'  The  hemolysis  he  attributes  to  direct  injury  of 
the  blood  in  its  passage  through  the  heated  area,  and  not  to  the  action 
of  poisons ;  this  is  very  possible,  since  red  corpuscles  fragment  after 
being  heated  to  52°,  and  may  be  seriously  impaired  functionally  at 
45°.     There  are  many  authors,  indeed,  who  consider  tlie  blood  changes 

T2Vircl)o\v's  Arch.,  1900    (183),  377. 

7:!  Kavcnna  and  ^Tiiiassian  ( IJcf.  in  Tiioc-honi.  C'(>Mtr..  1003  (IK  34S )  stato  tliat 
blood  heated  outside  tlie  body  to  .'ir)°-60°  is  toxic,  and  causes  the  same  anatomical 
chan<,'es  as  does  death  from  l)\irning,  vhicli  findinff  is  corroborated  bv  ireisted. 
Arch.  klin.  riiir.,  lOOfi    (70),  414. 

74  Arch.  klin.  f'hir.,  1005    (7r)),  S4.'i. 

75  Virehow's  Arch..  1005  (ISO),  3(i7:  Zeit.  f.  Ilyg.,  lOOG   (54).  419. 
75aZeit.  Tmmuniliit.,   1015    (23),  473. 


POISONS  PRODUCED  IN  BURNS  565 

the  chief  cause  of  death,  but  the  weight  of  evidence  is  iu  favor  of  the 
theory  of  the  development  of  toxic  substances  in  the  burned  skin. 

Kutscher  and  Ileyde  '"^  believe  methyl  guanidine  to  be  the  toxic 
substance  eliminated  in  the  urine,  stating  that  it  produces  effects  sim- 
ilar to  that  caused  by  injections  of  the  toxic  urine  from  burn  cases. 
These  symptoms  are  quite  similar  to  those  characteristic  of  anaphy- 
laxis, and  Heyde  states  that  small  burned  areas  sensitize  an  animal 
to  later  injections  of  extracts  of  burned  tissue.  He,  as  well  as  Vogt, 
are  therefore  inclined  to  believe  that  some  cases,  especially  those  dy- 
ing unexpectedly  12  or  13  days  after  the  burning,  may  be  the  result 
of  anaphylactic  reaction  to  proteins  made  of  foreign  character  by  the 
heat.'^  The  newer  observations  concerning  the  presence  of  toxic  sub- 
stances in  the  urine  during  anaphylactic  intoxication  are  in  harmony 
with  the  findings  in  burn  cases,"'''  although  the  identity  of  methyl 
guanidine  with  the  toxic  agent  is  questionable. 

Burn  Blisters. — The  contents  of  burn  blisters  resemble  the  fluid  of 
inflammatory  edemas  generally.  K.  Morner  ^^  found  5.031  per  cent, 
of  proteins,  which  included  1.359  per  cent,  of  globulin  and  0.011 
per  cent,  of  fibrin ;  there  was  also  present  a  substance  reducing  cop- 
per oxide,  but  no  pyrocateehin. 

76  Cent.  f.  Physiol.,   lOll    (25),  441. 

77  Heyde,  Med.  Klinik,  1912    (8),  263. 

77a  See  Pfeiflfer,  Zeit.  Immunitiit.,  1913   (18),  75. 
78Skand.  Arch.  Physiol.,  1895    (5),  272. 


CHAPTER    XIX 

GASTRO-INTESTINAL    "AUTOINTOXICATION"    AND 
RELATED  METABOLIC  DISTURBANCES 

Under  this  heading  are  commonly  included  all  intoxications  that 
can  be  ascribed  to  the  absorption  from  the  gastro-intestinal  tract  of 
toxic  substances  that  have  been  formed  within  its  contents,  either 
by  the  action  of  the  digestive  ferments  or  of  putrefactive  bacteria. 
The  propriety  of  considering  such  conditions  as  examples  of  auto- 
intoxication is  properly  questioned,  since  it  is  often  difficult  to  deter- 
mine whether  the  putrefaction  occurred  within  the  body,  or  had 
already  taken  place  in  the  food  before  it  was  eaten.  But  even  those 
who  would  limit  the  use  of  the  term  autointoxication  to  intoxication 
with  the  products  of  cellular  metabolism,  must  admit  the  possibility 
of  products  of  metabolism  reentering  the  blood  from  the  contents  of 
the  bowels  through  the  intestinal  wall,  since  the  bile,  and  perhaps 
also  the  intestinal  juice,  contain  excrementitious  substances  which 
may,  in  case  of  defective  fecal  elimination,  be  reabsorbed  into  the 
blood.  Therefore,  in  gastro-intestinal  disturbances  we  have  the  pos- 
sibility of  both  true  autointoxication  and  intoxication  by  putrefactive 
products  occurring  together  in  an  inseparable  way,  and  the  common 
inclusion  of  gastro-intestinal  intoxication  in  the  discussion  of  auto- 
intoxication would  seem  to  be  justifiable  as  well  as  expedient. 

The  sources  of  poisonous  substances  arising  in  the  gastro-intestinal 
tract  are  numerous.  They  may  be  formed  either  from  the  food-stuffs, 
or  from  the  secretions  and  excretions  of  the  body  that  enter  the  ali- 
mentary canal ;  and  they  may  be  formed  either  by  the  digestive  fer- 
ments or  by  the  bacteria  of  the  intestinal  contents.  Hence  the  number 
of  these  products  is  enormous,  and  we  are  by  no  means  sure  that  those 
that  have  yet  been  identified  include  the  most  important  or  most 
toxic.  To  classify  the  poisonous  substances  that  are  known  to  be 
formed  in  the  alimentary  canal,  and  which  might,  under  certain 
conditions,  cause  an  intoxication,  is  extremely  difficult,  because  of 
the  uncertainty  of  our  information;  but,  using  as  a  basis  the  sources 
of  the  substances,  they  may  be  classilied  as  follows:  ^ 

I.  The  con.stituents  of  the  digestive  secretions,  including  the  bile 
salts  and  pigments,  pepsin,  and  trypsin. 

1  ^r<)(lifie(l  from  Weintravul.  Er<rob.  all<r.  riitlml..  lSi)7  (4),  1,  wlio  gives  ex- 
hauHtive  discussion   iuid   hiliiin^rapliy   to  tluil   date. 

r)t;ti 


VONSTITUENTf^  <>l'  l>l<! i:^riV H  FLI  ll>S  '^^' 

11.  Products  of  >^"7^\^^V*'^'r^oses    peptones,  amino-acids. 
(ft)   From  proteins— proteoses    pM"^ 

(b)   From  fats-fatty  acids  and  ^dycerol. 
III.  Products  of  putrefaction  and  fermentation: 

(a)    From  protenis:  ,..,,-,:„„i„    (tyrosine,    phenylalanine, 

M^   T^rnm    the   aromatic    radicals    vk^^"*^'"  '    t       .-^  . 

(1)   J^rom   tne   a  skatole-carbonic    (or  in- 

,2^  ^^r'tlirfattv  acid  radieals-fatty  acids    (especially 
'^  butyric    nd  acetic),  acetone,  »>»™'"-- »'"'™:,t;'.'=:r 
1«„  dioxide    hvdrogen,  ma.sh-gas.     Also  pto.muiis .  ca 
;   rputresci„e%thylide„d,amine,  >soamylamn. 
(3)  Fvom    the    s«lpln,r-conta,nmg    rad.cals--H,S,    methyl 
'    '  mereaptan,  ethyl  mereaptan,  ethyl  sulphid. 

<^^  ^Xttncid- r  Mlowin,  havi,.,  --'-'«^™rc' 
acetic,  propionic,  butync,  vale"*™-  ^f-  »>1'»'^"'=- 
and  succinic;  also  acetone,  CO,,  t^ti,,  n,. 

'''  '^ XheTfatty  acids,  as  well  as  bntyric  acid;  also  gly- 
cerol.    From  lecithin-choline,  neurine,  and  muscarme- 

like  bodies. 

I     THE  CONSTITUENTS  OF  THE  DIGESTIVE  FLUIDS 

These  call  for  but  brief  -nsideration    for   althov^h  many  of  tbem 

are  known  to  be  toxic,  ff  .*-  -  - -''^»-  ''^^  j'l^,,,  ,,ypsin, 

intoxication  ctbe..  >» j'^™  ^«^tS  when  in^ect^d  experimentally 

especially  the  latter,  are  ucemtrL   y  on^ear  ever  to  pass 

:SiX»r::gen.  t&;"n  f:nbu;;;:  a:tion  „f .. .... 

"^Th/brsllts  are  also  toxic.  especia%  hemolytic,  ""*  tj- «-^ -;. 
reabsorbed  from  the  intestines  are  taken  ^^^  "'*»  '^^    ,  tTor  a  1 

^   1      Tiiw  T^vntpctive  arransrement  seems  to  be  sutncuuT  lui 
^^^^•^^^^^J^^Tirble^i'^ments  become  converted  into  urohihnogen 
emergencies.     Ihe  bile  pi^mt  ^  absorbed  and  eliminated  as 

through  reduction,   and   this  is  huge  >    ^^^^^'  ^^  ^  .^^^^^^^^^  ^ 

urnhiJin      Icterus  and  cholemia  do  not  seem  e^tI  \"  ^^  i 
WL  of  bile-pigments  and  bile  salts  from  the  nitestn.es. 


568  G ASTRO-INTESTINAL    ''AUTOINTOXICATION 


II.     PRODUCTS  OF  NORMAL  DIGESTION 

Proteoses  and  Peptones. — Under  normal  conditions,  these  are 
broken  uj)  in  the  intestinal  wall  into  the  amino-acids,  through  the 
agency  of  erepsin,  and  do  not  appear  in  the  blood  in  appreciable 
quantities.  To  be  sure,  certain  authors  claim  to  have  found  albumose 
in  nonnal  blood,  but  if  present  the  amounts  are  extremely  minute. 
In  conditions  in  which  ulceration  or  other  lesions  are  present  in  the 
gastro-intestinal  tract  it  is  possible  to  find  small  amounts  of  proteoses 
in  the  urine,  probably  absorbed  through  the  abnormal  areas,  but 
not  in  quantities  sufficient  to  account  for  any  appreciable  intoxication, 
although  proteoses  are  distinctly  toxic.  This  last  statement  has  been 
much  contested,  because  the  difficulty  of  purifying  proteoses  ob- 
tained from  ordinary  sources  has  left  open  the  possibility  that  such 
toxic  effects  as  have  been  observed  are  due  to  contaminating  sub- 
stances, and  not  to  the  proteoses  themselves.  More  recent  work,  how- 
ever, particularly  that  of  Underbill,-  Gibson  -^  and  Zunz,^  seems  to 
have  established  affirmatively  the  toxicity  of  proteoses,  whether  from 
animal  or  vegetable  proteins.  Besides  the  classical  effect  of  inhibiting 
the  coagulation  of  the  blood,  the  proteoses  have  a  lymphagogue  effect 
(Ileidenhain),'*  cause  a  marked  febrile  reaction,^'  and  in  doses  of  some 
size  are  fatal  to  experimental  animals  (rabbits  being  much  less  sus- 
ceptible than  dogs  and  many  other  animals).  Locally  they  cause  a 
mild  inflammatory  reaction,  which  is  followed  by  the  appearance  of 
much  connective-tissue  formation.**  Long  continued  injection  of  pro- 
teoses does  not  produce  visceral  lesions."'"^  The  careful  studies  of  Zunz 
show  that  intravenous  injection  of  hetero-albumose,  thio-albumose, 
deutero-albumose  and  proto-albumose  cause  a  rise  in  blood  pressure, 
but  large  doses  may  cause  a  fall  in  pressure ;  the  abiuret  products  of 

2Amer.   Jour.   Physiol.,    1903    (9),   ?A5;    Jour.    Biol.    Chcni.,    1015    (22),    443, 
(literature) . 
2a  Philippine  Jour.  Sei.,   1914    (9).  490. 

3  Arch,  intcrnat.  plivsiol.,  1911    (73),   110. 

4  F5ee  also  Nolf,  Arch,  internat  de  Diysiol.,   1906    (3).   343. 

5  Gibson  finds  that  carefully  purified  proteoses  liave  but  a  slight.  ])vrofrenio  ef- 
fect.     (Philippine  Jour.  Sci./l913    (8),  475.) 

6  In  a  paper  appearing  in  tlie  Transactions  of  the  Chicago  Pathological  Society, 
1903  (5),  240,  I  published  the  observation  that  repeated  injections  of  Witte's 
"peptone"  (which  consists  chiefly  of  proteoses)  into  rabbits  led  to  tlie  ]>roduction 
of  marked  cirrhosis  of  the  liver,  and  sujrgested  the  possil)ility  that  jiroteoses 
escaping  tlirougli  a  diseased  gastr'c  or  intestinal  wall  into  the  blood  might  be 
a  factor  in  tlie  production  of  cirrhos's  in  man.  Subse()uent.  observations,  how- 
ever, have  shown  tliat  rei)eated  injection  of  almost  any  foreign  protein  material 
(e.  g.,  emulsions  of  organs,  foreign  })lood.  etc.,  used  in  immunization  exjieriments) 
will  cause  a  similar  cirrliosis  in  ral)bits,  which  animals,  indeed,  often  spon- 
taneously sliow  this  condition  when  apparently  otherwise  normal.  "Peptone"  in- 
jections in  dogs  and  guinea-pigs  have  failed  to  cause  a  similar  cirrhosis,  and  lu'nce 
the  value  of  tliese  and  all  other  rabbit.  exi)eriments  on  cirrliosis  of  the  liver  is 
very  questionable:  however,  the  possibility  of  the  correctness  of  the  original  con- 
clusions still  remains  open. 

flaWoolley  et  al.,  Jour.  Exp.  Med.,  1915   (22),  114. 


PRODUCTS  OF  .Yr>AM/.l/>  D/dESTION  569 

tryptie  digestion  are  iiiurli  more  actively  depi'essor  than  tlie  allnmioscs. 
As  a  general  rule,  however,  it  has  been  observed  that  the  first  products 
of  protein  hj-drolysis  are  the  most  toxic,  and  with  further  cleavage  the 
toxicity  lessens  and  finally  disappears,  as  sliown  especially  in  the 
studies  on  ana])hy]axis  and  anaphylatoxin  formation.""  Thus  AVolf  ^ 
found  that  the  amino-acids  do  not  cause  a  fall  of  blood  pressure,  nor 
do  polypeptids.^ 

Most  of  the  attempts  to  obtain  antibodies  for  the  products  of  pro- 
tein hydrolysis  have  failed,  for  it  seems  to  require  an  intact  protein 
molecule  to  act  as  antigen.  The  most  authentic  positive  results  so  far 
reported  are  those  of  Zunz,"  who  claims  that  with  the  higher  cleavage 
products,  the  ''primary  proteoses"  from  fibrin  hydrolysis,  he  secured 
some  degree  of  anaphylactic  reaction.  This  work  has  failed  of  con- 
firmation and  with  the  cleavage  products  of  egg  albumen  Wells  ^*' 
obtained  no  positive  results. 

"Albumosuria."  ""^ — If  proteoses  enter  the  blood  stream  they  ap- 
pear in  large  part  in  the  urine,  indicating  that  the  tissues  do  not  read- 
ily utilize  them  in  this  form."  Consequently,  when  proteoses  are  pro- 
duced in  considerable  amounts  by  autolysis  of  pathological  tissues 
they  appear  in  the  urine,  and  their  presence  is  considered  to  be  of  di- 
agnostic value. ^^  True  peptone  seems  rarel.y,  and  according  to  manj' 
observers  never,  to  appear  in  the  urine.  But  in  view  of  the  observa- 
tions that  polypeptids  often  appear  in  the  urine,^-''  it  is  probable  that 
true  peptones  also  do.  Albumoses,  therefore,  may  be  found  in  the 
urine  whenever  any  considerable  amount  of  tissue  or  exudate  is  being 
autolyzed  and  absorbed,  and  it  has  been  found  in  the  following  con- 
ditions: Suppuration  of  all  kinds;  resolution  of  pneumonia;  involu- 
tion of  the  puerperal  uterus;  carcinoma  (two-thirds  of  all  cases — Ury 
and  Lilienthal ) ,  and  other  malignant  growths ;  febrile  conditions  vrith 
tissue  destruction  (37.5  per  cent,  of  all  eases,  ]\[orawitz  and 
Dietschy)  :  "  acute  yellow  atrophy,  phosphorus  poisoning,  and  eclamp- 
sia; leukemia,  especially  under  .r-ray  treatment;  absorption  of  simple 
and  inflammatory  exudates;  ulcerating  pulmonary  tuberculosis,^*  and 

6b  The  statement  of  v.  Knaffl-Lenz  (Arch.  exp.  Path.  u.  Pharm.,  1913  (73), 
292)  that  the  toxicity  of  the  cleavage  products  varies  directly  with  their  trypto- 
phane content  could  not  be  corroborated  bv  Underbill  and  Hendrix,  Jour.  Biol. 
Cheni.,  1915   (22).  443. 

7. Tour,   of   Phvsiol.,    100.5    (32),    171. 

8  Halliburton, \biVZ.,  190.5    (32),  174. 

9  Bull.  Acad.  Roval  de  M^^d.  Belgique,  Mav  27,   1911. 

10  Jour.  Infect.  Dis.,  1900    (6),  506. 

lOaGood  critical  review  given  by  Pollak.  Zeit.  exp.  ^led..  1014    (2).  314. 

11  They  may  be  partly  liydrolyzed  into  smaller  complexes,  however,  primary  pro- 
teoses being  partly  changed  to  deutero-proteoses,  and  the  latter  partly  to  peptones 
(Chittenden,  ^lendel,  and  Henderson,  Amcr.  Jour.  Phvsiol.,  1S99    (2),  142). 

12  See  Yarrow.  Amer.  Med.,  1903  (5),  452;  Try  and  Lilientlial,  Arch.  f.  Ver- 
dauungskr.,  1905   (11),  72;  Senator,  International  Clinics,  1905   (4,  series  14),  85. 

i2aChodat  and  Kummer,  Biochem.  Zeit..  1914    (05),  392. 

13  Arch.  f.  exp.  Path.  u.  Pharm.,  1905    (54),  88. 

14  See  Parkinson,  Practitioner,  190G   (76),  219. 


570  GASTROIXTESTIXAL    'AUTOIXTOXICaTIOX" 

after  tuberculin  reactions  ( Deist ).^^  Albumosuria  is  present  in  small- 
pox and  may  serve  in  differential  diagnosis.' ■'^''  In  ulcerative  condi- 
tions of  the  alimentary'  canal  albumoses  may  be  absorbed  unchanged 
and  cause  alimentary  albumosuria.  The  normal  kidney  seems  to  be 
impermeable  to  the  small  amounts  of  proteose  that  may  be  present  nor- 
mally in  the  blood,  or  even  after  large  oral  ingestion  of  proteoses,  but 
in  parenchymatous  nephritis  it  may  escape  in  the  urine  (Henderson/** 
Pollak^"^).' 

It  is  possi])le  that  some  of  the  symptoms  of  these  conditions  are 
due  to  intoxication  with  proteoses,  for  0.07  to  0.1  gram  deutero-albu- 
mose  will  cause  a  febrile  reaction  in  a  healthy  man/'  but  probably 
their  amount  is  usually  too  small  to  cause  appreciable  etfects.^^  It  is 
well  known,  however,  that  the  characteristic  rise  of  temperature  fol- 
lowing tlie  injection  of  tuberculin  into  tuberculous  individuals  is  also 
produced  if  minute  quantities  of  proteose  solutions  are  injected  in 
place  of  tuberculin ;  therefore,  proteoses  arising  from  autolysis  in  tu- 
berculosis may  be  of  importance  in  causing  fever  and  other  symp- 
toms.^^  Tuberculous  animals  are  said  to  succumb  to  a  much  smaller 
dose  of  deutero-albumose  than  normal  auimals.^®^ 

The  so-called  "Bence- Jones  albumose"  that  appears  in  the  urine 
of  patients  with  multiple  bone-marrow  tumors  is  not  a  true  albumose, 
but  is  more  closely  related  to  the  simple  proteins,  and  is  discussed 
under  the  head  of  "Chemistry  of  Tumors." 

III.     PRODUCTS  OF  PUTREFACTION  AND  FERMENTATION  -° 

AVe  may  perhaps  gain  some  appreciation  of  the  enormous  amount 
of  bacterial  action  that  goes  on  in  the  normal  intestinal  digestive 
processes  by  considering  the  fact  that  as  much  as  one-third  of  the  total 
weight  of  the  solids  of  normal  feces  may  consist  of  bacteria  (Stras- 
burger),  their  proportion  being  increased  in  diarrheal  disorders  and 
decreased  in  constipation.  They  attack  all  food-stuff's,  and  among 
the  decomposition-products  formed  through  their  activity  are  un- 
doubtedly many  of  considerable  toxicity.  ]\Iost  of  the  products  of  in- 
testinal putrefaction  that  have  as  yet  been  isolated  are,  however,  not 
extremely  poisonous ;  but  many  of  them  are  toxic  to  some  degree,  and 
their  long-continued  absorption  may  well  lead  to  serious  disturbances. 

15  Boilr.  z.  kliii.  'I'lilH-rk..  I!tl2   (23),  .147. 

isa  Primavora,  Cay..  Int.  Med.  o  Cliir.,  IIU.3,  No.  10. 

i«Laiu-pt,  :Mar.  6,  IflOO. 

17  See  :\rattlies.  Arch,  exper.  Patli.  u.   Pliarni..  ISO.")    CM)).  4:i7. 

1**  In  a  series  of  uni)ul»lislu'd  experiments  I  was  nnable  to  cause  amylnid  de- 
feneration in  rabbits  by  ]>r()tracted  intoxication  with  proteose  solutions. 

i!»  Simon,  .\rch.  ex]).  :Med..  190:?  (4!t),  44!>.  Concernini:  relation  of  tuberculin 
to  proteoses  see  review  liy  -lolles  in  Ott's  "C'hemisi'he  l\itliol.  der  Tubcrculose." 

i!ia  Kirchlieim  aiid  Tuczek.  Arcli.  exj).  Path.  u.  I'harm..  1014    (77).  ."^87. 

2"  Complete  bibiiy^rajjliv  jriven  in  the  resumf'  on  "Intestinal  Putrefaction"  bv 
Gerhardt,  Erpebnisse  der  Physiol.,  1004  (III.  Abt.  1).  107.  Chemistry  of  Putre- 
faction is  reviewed  by  Ellinjjer,  ibid.,  1007    ( ti ) ,  20. 


I'h'ODl  t-TS  or  rUTREFACTION  AND  FERMENTATWX  571 

Considering-  tlu'iu  lirst  according  to  their  origin  and  chemical  nature, 
we  take  up  iirst  the  products  of: 

A.     PROTEIN  PUTREFACTION 

(1)   SUBSTANCES  DERIVED  FROM  THE  AROMATIC  RADICALS   OF 
THE   PROTEIN   MOLECULE 

In  the  protein  molecule  are  contained  the  following  amino-acids 
with  an  aromatic  nucleus : 

Tyrosine,  Ho/      y-CH^—  CH  —  COOH 

riu'iiylalaniiu',  <^       \  cH.- —  CH  —  COOH 

NH2 
Ti  yptopliane,  /      N_  C  —  CH2  —  CH  —  COOH 

\         CH 

V 

H 

In  the  intestinal  contents  have  been  found  a  number  of  substances 
that  are  undoubtedly  derived  from  these  aromatic  radicals.  They 
are  (1)  phenol,  . — . 

which  is  formed  in  small  quantities,   presumabl}^  from  tyrosine,   as 
also  is  the  closely  related  (2)  paracresol, 

and  also  (3)  para-oxyphenyl  acetic  acid, 

HO  /      \  CH,  —  COOH 
and  (4)  para-oxyphenyl-propionic  acid, 

HO  /      N  CH,  —  CH..  —  COOH 


From  the  tryptophane  are  formed  numerous  important  substances, 

as  follows: 

NH2 


_  C  —  CH2  —  CH  —  COOH 

\/ '" 

XH 
(tryptophane) 


readily  yields,  through  splitting  off  the  NIT,  group  and  addition  of 
H,  iiidole  propionic  acid  (formerly  incorrectly  called  sl-atole  acetic 
acid),  which  is 


572  GASTROINTESTINAL    "AUTOINTOXICATION" 

CH=  —  CH2  —  COOH 
/ 

C 
\^ 

I        CII 

I  / 

NI[ 

and  from  which  in  turn  may  readily  be  formed  indole  acetic  acid 
(erroneously  called  skatole  carhoxylic  acid),  which  is 

0H=  —  COOH 

CH 
I      / 
NH 

Both  of  these  substances  have  been  formed  in  the  intestinal  contents. 
From  these  substances  are  formed  the  better  known  skatole, 

cm 
/ 
c 
,    W 

CH 

I      /  . 

NH 

and  indole,  / — v 

<        >CH 

CH 

NH 

In  dogs,  but  not  in  man,  kynurcnic  acid.  / — \ 

<        >—  C.OH 
^-/      \^ 

C.COOH 

N  =  CH 
is  also  formed  from  tryptophane.-^ 

The  greatest  interest  concerning  these  bodies  arises  from  the  fact 
that  after  they  are  absorbed  from  the  intestine  they  become  combined 
with  sulphuric  or  glycuronic  acid,  and  are  excreted  in  the  urine  as 
salts  of  these  acids ;  consequently  the  amount  of  sulphuric  acid  ap- 
pearing in  the  urine  in  such  organic  combination  ("ethereal  sul- 
phuric acid")  is  considered  as  an  index  of  the  amount  of  intestinal 
putrefaction.  In  the  case  of  indole  and  skatole,  which  have  no 
hydroxyl  group,  a  preliminary  oxidation  occure,  whereby  inHole  is 
converted  into  indoxyl, 

>C  —  OH 

I        CH 

I  / 

NH 


and  skatole  into  skatoxyl. 


\ 


C  —  ClJs 

roH 
/ 

XH 


21  See  Ellingcr,   Zcit.   pliysiol.  CIiciii.,   1!)04    (43),  325. 


PRODUCTH  OF  PUTREFACTION  A\D  FEIIMENTATIOX  573 

and   they  are  thou  combined  with  sulphuric  or  glycuronie   acid,  as 
follows : 


_  c  — ;  OH  +  H :  0  —  SO,  —  OK  =  <       >  _  c  —  o  —  SO2  —  ok 
N\  ■• '  V_y     N\ 

\  CH  \  pB.      (indican) 

HN  -\"H 

B}^  far  the  greater  part  of  these  aromatic  substances,  when  ex- 
creted in  the  urine,  is  combined  with  sulphuric  acid,  and  but  a  small 
part  with  glycuronie  acid ;  but  in  case  the  amount  of  sulphuric  acid 
available  is  too  small  to  combine  with  all  the  aromatic  radicals  enter- 
ing the  blood,  a  large  amount  of  the  glycuronie  acid  compound  ap- 
pears in  the  urine  (e.  g.,  after  therapeutic  administration  of  phenol, 
cresol,  thymol,  camphor,  etc.)-  Both  the  preliminary  oxidation  and 
the  combining  with  acids  seem  to  occur  chiefly  in  the  liver,  this 
process  constituting  one  of  the  most  important  of  the  many  pro- 
tective offices  of  that  organ,  since  the  resulting  compounds  are  much 
less  toxic  than  are  the  original  substances.-^''  Herter  and  Wakeman  -- 
have  shown  that  living  cells  have  the  power  of  acting  upon  indole 
and  phenol  (and  presumably  upon  the  rest  of  this  group)  in  such  a 
way  that  they  cannot  be  recovered  by  distillation.  Most  active  in  this 
respect  is  the  liver,  then  in  order  come  kidney,  muscle,  blood,  and 
brain.  The  change  seems  to  be  a  loose  chemical  combination  with  the 
protoplasm  of  the  cells,  and  the  power  of  the  tissues  to  bring  about 
this  combination  is  not  greatly,  decreased  by  serious  pathological 
changes  in  the  organs  (e.  g.,  ricin  poisoning).-^ 

Indole. — This  is  probably  the  most  important  member  of  this  group 
of  substances,  the  striking  color  of  its  derivatives  making  its  detec- 
tion in  the  urine  easy,  so  that  it  is  generally  used  as  the  most  available 
index  of  the  amount  of  putrefaction  that  is  occurring  in  the  intes- 
tines.^* The  greatest  quantities  are  found  when  intestinal  putre- 
faction is  marked,  especially  in  intestinal  obstruction  involving  the 
small  intestine ;  obstruction  of  the  large  intestine,  as  Jaife  first  dem- 
onstrated, does  not  cause  marked  indicanuria  unless  the  stagnation 
involves  the  ileum,  as  it  may  in  the  latter  stages  of  obstruction.  With 
marked  impairment  of  renal  function  indican  may  accumulate  in  the 
blood  (see  Uremia).  There  can  be  no  question  that  the  indican  of  the 
urine  is  derived,  at  least  in  part,  from  the  indole  formed  in  the  intes- 
tine, for  administration  of  indole  by  mouth  to  either  animals  or  man 
causes  a  considerable  increase  in  the  indican  present  in  the  urine ;  how- 
ever, but  40  to  60  per  cent,  can  be  recovered  in  this  way,  the  rest  ap- 
parently being  oxidized  ^0  other  compounds,  part  of  which  may  also 

21a  Metchnikoff  insisted  that  these  sulfo-compounds  still  retain  considerable 
toxicity.      (Ann.  Inst.  Pasteur,  1014    (27),  80."^). 

22  .Tour.  Exper.  :^red..  ISOt)    (4),  307. 

23  For  further  discussion  of  this  topic,  see  "Chemical  Defences  against  Poisons 
of  Known  Composition,"  Chapter  ix. 

24  See  Houghton,  Amer.  Jour.  Med.  Sci.,  1908  (135),  567. 


574  GASTRO-INTESTIXAL    '■AVTOiyTOXICATlOX" 

aiipear  in  tlie  urine. -■'  Whether  i)art  of  tlie  urinary  indiean  is  derived 
from  tryptophane  liberated  during  intracellular  protein  metabolism, 
and  not  from  intestinal  putrefaction,  has  long  been  a  disputed  point 
among  physiological  chemists.-"  The  demonstration  hj  Ellinger  and 
Gentzen  -'  that  try]ito])hane,  when  fed  or  injected  su1)cutaneously, 
causes  no  increase  in  uriiuiry  indican,  whereas  its  injection  into  the 
cecum  causes  much  indicanuria,  would  indicate  that  indole  is  formed 
from  tryptophane  only  through  putrefaction,  and  not  in  cellular 
nietabolisiu.  Othei-  expei'imeuts  support  the  same  view.-*^  However, 
it  is  possible  that  part  of  the  indican  present  in  the  urine  during 
conditions  associated  with  gangrene,  putrid  cancers,  putrid  placentas, 
or  puti'id  purulent  exudates,  may  be  derived  from  these  decomposing 
materials.  The  statement  that  indicanuria  is  of  significance  in  in- 
sanity could  not  be  substantiated  by  Borden,-"  who  used  quantitative 
methods  and  careful  controls.  A  large  proportion  of  the  data  and 
conclusions  in  the  literature  concerning  indicanuria  are  valueless  be- 
cause of  improper  or  inadequate  methods. 

Probably  the  chief  agent  in  the  formation  of  indole  in  the  intes- 
tines and  in  putrid  tissues  is  the  colon  bacillus,  which,  as  is  well 
known,  produces  indole  in  ordinary  culture-media. 

Toxicity  of  Indole. — Although  the  toxicity  of  indole  seems  to  be 
relatively  slight,  and  this  toxicity  is  further  reduced  by  the  conver- 
sion of  indole  into  indoxyl  and  indican,  yet  Herter  ^^  found  that  ad- 
ministration to  healthy  men  of  indole  in  quantities  of  0.025  to  2  grams 
per  day  caused  frontal  headache,  irritability,  insomnia,  and  confu- 
sion ;  the  continued  absorption  of  enough  indole  to  cause  a  constant 
strong  reaction  for  indican  in  the  urine  is  sutificient  to  cause  neuras- 
thenic symptoms.  Long-continued  injection  of  indole  leads/to  hyper- 
trophy of  the  adrenal  medulla  and  slight  interstitial  changes  in  the 
kidneys,^^  but  the  reputed  responsibility  of  indole  for  arteriosclerosis 
is  most  doubtful. ^^^  Lee  ^-  has  also  demonstrated  that  iiulole.  skatole, 
and  methyl  mercaptan  cause  muscles  to  react  to  stimuli  like  fatigued 
muscles.  Normal  urine  contains  but  about  12  milligrams  of  indican 
per  day,  which  amount  is  so  insigiiificant  in  proportion  to  the  above- 
mentioned  doses  that  were  found  necessary  to  produce  symptoms, 
that   we   may   well    doubt   the   occurrence   of   noticeable   intoxication 

-^'  If  frt'latin  is  subaiitutod  for  proteins  in  tlic  diotary,  indican  is  not  oxrrcted, 
because  {gelatin  does  not  contain  tryptophane  (Underliill,  Amer.  .lour.  Piivsiol., 
1!KM   (VI),  17(i). 

20  Literature  by  Gerliardt,  Erfrel).  der  I'liysiol.,  1!)(I4    (  11  [.  Abt.  T),  LSI. 

27  TL)fineister's  Beitr.,   1903    (4),   17L 

2sSee  Scliolz.  Zeit.  physiol.  Clieni..  1!)0.3  (.'JS),  fiL'?;  ITnderhill.  Jor.  cit.  Slierwin 
and  Hawk  found  an  absence  of  indican  in  the  urine  in  tlie  latter  part  of  a  long 
fast    (ISiocliem.  Bull.,   1914    (3),  410). 

2ft  Jour.    Uiol.   Cheni.,   1907    (2),  r)75. 

30  X,.\v   York  Med.  dour.,   1S9S    (6S),  89. 

31  W.xdlev  and  Xewburgb,  .lour.  Amer.  IMed.   Assoc.   1911    (oO),   1796. 
siaSee  Steenliuis,   Folia   Mikroliiol..   1915    (3).  7(>. 

32  Jour.  Amer.    Med.   .\ssoc.,    190()    (40),    149!t. 


I'noitt  <Ts  or  I'l  Th'inAci  /(>\    \\i)  ji:inii:\T  \'H()\  575 

from  tliis  siil)stiiii('t'  iiiidcr  ordinary  coinlitious.  Nesbitt  "•'  states  that 
twenty  tiuics  as  iiindi  iiulok'  or  skatole  as  are  excreted  daily  by  an 
adult  man  may  be  injected  into  the  ju<rular  vein  of  a  dog  of  four 
kilos  without  causing  appreciable  effects.  Richards  and  Ilowland, 
liowever,  have  demonstrated  the  possibility  that  defective  oxidation  of 
substances  of  this  group  nuiy  permit  of  intoxication.^' 

Other  Aromatic  Compounds. — Skatole  seems  to  accompany  indole 
in  small  amounts,  but  ai)pai"ently  in  no  constant  quantitative  rela- 
tion. Herter  ^^  states  that  skatole  is  fonned  under  entirely  different 
conditions  from  indole,  and  that  B.  coli  does  not  produce  skatole. 
It  is  not  always  present  in  the  contents  of  the  large  intestines  of 
healthy  persons,  and  seems  to  be  formed  later  than  indole. 

Indole-acetic  acid  appears  in  the  normal  urine  in  extremely  minute 
quantities,  and  is  increased  in  the  same  conditions  as  skatole.  It  is 
the  mother  substance  of  urorosein,  and  can  be  found  in  the  intestines 
of  patients  who  show  this  substance  in  the  urine  ( Herter ).^'^  Ross  ^*^'^ 
found  indole-acetic  acid  in  the  urine  in  21  per  cent,  of  normal  persons, 
and  in  48  per  cent,  of  dementia  precox  cases,  and  obtained  evidence  in 
favor  of  an  endogenous  origin  in  two  cases  studied  especially  to  de- 
termine this  point. 

Phenol  ^"^  appears  in  the  urine  normally  in  very  minute  quantities — 
from  0.005  to  0.07  grams  per  day,  according  to  various  observers. 
These  figures  are  undoubtedly  too  low,  for  Folin  and  Denis  ^^''^  found 
the  total  excretion  of  phenols  to  be  from  0.2  to  0.4  gm.  per  day. 
Much  more  is  undoubtedly  formed  in  the  intestines,  for  but  a  small 
fraction  of  phenol  given  by  mouth  (2  to  3  per  cent.,  according  to 
]\Iunk)  appears  in  the  urine  as  a  sulphuric-acid  compound;  part  of 
the  rest  is  oxidized  to  hydrochinon  and  pyrocatechin,  CcH^(OH).,, 
and  eliminated  as  ethereal  sulphates.  These  sulphates,  although  dis- 
tinctly toxic,  are  much  less  so  than  the  phenol  itself  (Metchnikoft').^'' 
(^ontrary  to  prevailing  ideas,  Folin  and  Denis  found  the  greater  part 
of  the  phenols  to  be  excreted  uncombined.  The  largest  quantities  are 
found  in  the  same  conditions  as  indican,  except,  of  course,  in  ' '  carbolic- 
acid"  poisoning,  when  the  amounts  may  be  so  great  that  practically 
all  the  sulphuric  acid  in  the  urine  is  in  this  organic  combination,  much 
of  the  phenol  under  these  conditions  being  also  combined  with  gly- 
curonic  acid."''  This  pairing  is  accomplished  chiefly  in  the  liver 
(Dubin).  A  small  amount  of  urinary  phenol  maj^  be  of  endogenous 
origin.^*'' 

33  .Jour.  Exper.  :Mcc1.,   189!)    (4),  5. 

34  See  note  in  Science,  1006   (24),  970. 

3 s  Jour.  Biol.  Chem.,  1008    (4),  101;  general  discussion. 

3«  Jour.  Biol.  Chem.,   1008    (4),  253. 

36a  Arch.  Int.  Med.,  1013    (12),  112  and  231. 

3Cb  Literature  given  bv  Dubin,  Jour.  Biol.  Chem.,  1016    (26),  60. 

36c- .Jour.  Biol.  Chem.."l01,T    (22).  300. 

37  Ann.  Inst.  Pasteur,   1010    (24),  7.')0. 

38  See  the  oltservations  of  Wolilgemutli  and  of  Blumentlial  (Biochem.  Zeit- 
schrift,  1906   (1).  134),  on  tlio  detoxication  of  Ivsol  and  similar  poisons. 

38a  Moore,  Amer.  .Jour.  Dis.  Child.,  1917   (13),' 15. 


576  GASTROINTESTINAL    "AUTOINTOXICATION" 

Cresol  (chiefly  paracresol),  para-oxyphenyl  acetic  acid,  and  para- 
oxyphenyl  propionic  acid  appear  under  similar  conditions,  except 
that  the  two  oxy-acids  are  possibly  also  formed  within  the  body 
through  cellular  metabolism,  as  they  have  been  found  present  in  the 
urine  independent  of  intestinal  putrefaction.  Paracresol  is  quanti- 
tatively the  most  important  of  the  urinary  phenols,  and  long  continued 
feeding  produces  no  noticeable  effects  in  rabbits."^'*''  Probably  part  of 
the  benzoic  acid  that  appears  in  the  urine  combined  with  glycocoll,  as 
hippuric  acid,  is  derived  from  intestinal  putrefaction.^'' 

THE  PRESSOR  BASES  40 

Among  the  products  of  protein  putrefaction  are  several  amines  that 
have  marked  power  to  stimulate  the  sympathetic  nervous  system  and 
thus  to  raise  the  blood  pressure,  hence  resembling  epinephrine  physio- 
logically as  well  as  chemicall.y.  Since  increase  in  blood  pressure  is  so 
important  a  condition  in  clinical  medicine  these  substances  are  of 
much  significance.  The  most  important  of  the  pressure-raising  bases 
are,  in  the  order  of  their  pressor  effect : — 

Phenyl-ethylamine  C„H,.CH,.CH,.XH„ 

derived  from  phenylalanine  C,H,.CH,.CHNH,.COOH. 

Para-hj'droxy-phenjd-ethylamine  ( t^ramine )     Oil . C Jl^ . CIL . CHj . XH, 

derived  from  tyrosine  OH.CeH,.CH,.CHXH,.COOH. 

Its  relation  to  epinephrine  is  seen  on  comparing  with  the  structural 
formula  of  the  latter  ( OH )  ,CeH3 . CHOH . CH.NH . CH3 

.XH  — CH 
Beta-iminazolyl-ethyl'amine  (histamine)  CH^ 


^X     —  C     —  CH„:CH,.XH, 


/XH  —  CH 
derived  from  histidine  CH^  || 

\\X     —  C     —  CH, .  CHXH„ .  COOH 

It  will  be  observed  that  these  are  all  amines  derived  from  the  cyclic 
amino-aeids  of  proteins  by  the  process  of  decarboxylization  (loss  of 
COo),  a  process  which  occurs  only  under  anaerobic  conditions.  The 
straight  chain  amines  are  much  less  active  ])hysiologically.  These  sub- 
stances have  all  been  found  in  puti'id  protein  materials,  produced  by 
the  action  of  anaerobic  bacteria,  and  possibly  they  are  formed  in  the 
intestines.  Para-hydroxy-phenyl-ethylamine  is  also  one  of  the  active 
constituents  of  ergot,  and  has  been  found  in  the  salivary  gland  of 
cephalopods,  where  it  functions  as  a  venom  in  parcilyzing  the  prey. 
Beta-iniiiiazolyl-ethylamine  is  also  })resent  in  ergot. 

These  substances  are  all  detoxicated  by  the  liver,  and  hence  have  lit- 

3«b  Denny  and  Frothingliam,  Jour.  Med.  Res.,  1914   (31),  277. 

30  See  Prager,  Med.  Xews,  1005  (SG),  1025;  Magnus-Levy,  Miincli.  med.  Woch., 
1905    (52),  2168. 

40  Full  bihliography  given  l)y  Hargor,  "The  Simph'r  Xatural  Bases,"  Mono- 
graphs on  Biochemistry,  London,  1914. 


ALKM'TOM  h'lA  577 

tie  effect  when  <i'iveii  by  inouth/'"'  'J'licii-  detoxication  is  accomplished 
by  deainiiii/.ation  and  oxidatioii,  the  n'sultinjr  (•ar])()xylic  acids  being 
excreted  or  burned.  Therefore  it  is  not  certain  whether  pressor  bases 
formed  in  the  intestines  have  any  pathological  effect,  but  it  is  quite 
possible  that  outside  the  portal  territory  various  infections  may  give 
rise  to  pressor  bases  which  enter  the  general  circulation  directly  and 
escape  the  defensive  mechanism  of  the  liver.  They  may  also  cause 
local  effects  in  the  tissues  where  they  are  formed,  e.  g.,  the  bronchi. 
In  some  respects  their  effects  resemble  those  of  anaphylactic  intoxica- 
tion, and  as  the  latter  apparently  results  from  toxic  products  of  protein 
cleavage  the  possibility  that  here  too  pressor  bases  are  concerned  at 
once  presents  itself,  but  as  yet  the  relation  is  undetermined  (see  Ana- 
phylaxis, Chap.  vii).  The  resemblance  is  especially  seen  in  the  pro- 
found effect  on  the  bronchial  musculature,  which  can  be  thrown  into 
strong  contraction,  especially  by  beta-iminazolyl-ethylamine  which  in 
0.5  mg.  doses  kills  guinea  pigs  from  asphyxia,  with  distended  lungs  as 
in  fatal  anaphylaxis;  also  it  causes  a  similar  fall  in  temperature,  but 
does  not  render  the  blood  incoagulable.  Another  point  of  similarity 
is  the  severe  local  urticaria  when  weak  solutions  (1-1000)  of  histamine 
are  placed  on  a  scarified  area  of  skin,*"*"  recalling  vividly  the  fact  that 
urticarial  eruptions  are  conspicuous  in  some  tj'pes  of  anaphjdactic 
reactions.*^'^ 

ALKAPTONURIA  ii 
Alkaptonuria  may  be  appropriately  considered  in  this  connection, 
since  it  depends  on  an  abnormal  metabolism  of  the  aromatic  groups, 
tyrosine  and  phenylalanine,  which  are,  partly  at  least,  split  out  of 
the  protein  molecule  in  the  intestine.  This  condition  is  character- 
ized by  the  tendency  of  the  urine  to  turn  dark  on  exposure  to  air. 
due  to  the  presence  in  it  of  homogentisic  acid.*-  Homogentisic  acid 
has  been  found  in  the  blood  but  not  in  the  feces  of  alkaptonurics,  and 
the  urine  shows  no  other  deviations  from  the  normal  except  a  slight 
increase  in  ammonia,  with  which  the  acid  is  combined.  It  is  of  rare 
occui-rence,  persists  throughout  life  with  but  little  apparent  effect 
upon  the  health  of  the  individual,  and  is  often  hereditary,  being 
grouped  by  Garrod  along  with  cystinuria,  pentosuria  and  albinism 
as  a  "chemical  malformation"  of  hereditarv^  origin.*^     The  relation 

40b  See  Guggenheim  and  Loeffler,  Biochm.  Zeit.,  inifi    {12),  32.). 

■toc  Eppingcr  and  Outtmann.  Zeit.  klin.  Med.,   101.3    (78),  399. 

4id  Sollniann  lias  found  that  several  other  amines  give  urticarial  reaetions 
(Jour,  riiarni..  1917    (9),  301). 

*i  Resume  and  literature  by  Falta,  Biochem.  Centralhlatt.  1004  (3),  174.  and 
Deut.  Arcli.  klin.  Med.,  1904  (81),  231;  Garrod,  '-Inborn  Errors  of  [Metabolism," 
Oxford  :Med.  Publications.  1000;  also  Laneet,  .Tulv,  1008;  Frommherz,  Bioehem. 
Centr..   1008    (8),   1. 

■42  It  should  be  mentioned  that  hiidorhinon.  when  present  in  the  urine  (usually 
after  ingestion  of  large  quantities  of  phenol),  may  also  turn  dark  on  exposure 
to  air;  and  melanin  may  be  excreted  as  a  chromogen  which  turns  dark  on  ex- 
posure, by  patients  with  melanotic  tumors  or  ochronosis    {q.   v.). 

•13  Alkaptonurics  may  give  a  positive  WasserniauTi  reaction   without   otlu-.-  evi- 


578  GASTR0-INTE8TIKAL    "AUTOIXrOXICATIOy" 

of  these  aromatic  bodies  to  the  aromatic  constituents  of  the  proteins 
is  best  shown  by  comparing  their  structural  formula3 : 

rhenylalanine,  /^     \  CIL  —  CHXH,  —  COOH. 


Tyrosine,  HO—/         >  CIL  — CHXH„  —  COOH. 


OH 
Uroleucic  aci(i,->4  /     \  CH„  —  CHOH  —  COOH. 

HO 

OH 
Homogcntisic  acid,  /^      \  CK,  —  COOH. 

HO 
Apparently  the  condition  depends  upon  an  abnormality  in  the  in- 
termediary metabolism,  and  not  upon  an  abnormal  formatiou  of  homo- 
urentisie  acid  through  intestinal  putrefaction,  as  was  at  first  believed. 
Alkaptonuria  is  never  observed  in  slight  degrees;  if  there  is  any 
homogcntisic  acid  in  the  urine  at  all  it  is  there  in  large  amounts  (4-5 
grams  per  day),  depending  on  the  diet,  for  when  the  error  in  metabo- 
lism is  present  at  all  it  is  complete.  On  a  mixed  diet  the  ratio  of 
homogcntisic  acid  to  nitrogen  in  the  uriue  is  40^5  to  100.  The  pre- 
vailing idea  has  been  that  the  abnormality  consists  not  in  the  excessive 
formation  of  homogcntisic  acid,  but  in  a  lack  of  ability  on  the  part 
of  the  alkaptonurie  individual  to  split  open  the  benzene  ring.  It  is 
generally  stated  that  tyrosine  and  phenylalanine  first  suffer  a  split- 
ting out  of  the  nitrogen  radical  from  the  alanine  side-chain,  and  then 
are  oxidized  into  homogcntisic  acid,  following  which  changes  comes  a 
disintegration  of  the  benzene  ring,  with  subsequent  complete  oxida- 
tion. On  this  basis  the  alkaptouuric  accomplishes  the  conversion  into 
the  ox3^-acid,  but  the  process  stops  there.  Wakeman  and  Dakin,*^ 
hoAvever,  have  obtained  evidence  that  in  the  normal  oxidation  of  tyro- 
sine and  phenylalanine,  homogcntisic  acid  is  not  an  intermediary 
product,  and  Dakin  statop  that  the  alkaptonurie  can  destroy  simple 
derivatives  of  plienylalanine  and  tyrosine,  provided  their  structure 
is  such  that  the  formation  of  substances  of  tlu^  type  of  homogentisic 
acid  is  precluded.  He  believes  that  in  alkaptonuria  there  is  abnoi'mal 
formation  of  homogentisic  acid  as  well  as  a  failure  to  destroy  it  when 
formed.     On  the  other  hand,  Abderhalden  '**'  has  been  able  to  cause 

dence  of   syphilis,   and   in   one  case  this   reaction   disappeared    wlien    the   ]>atient 
was  given  large  amounts  of  tyrosine   (Siiderliergh,  Nord.  Med.  Arkiv.,  IHI.")   (48), 

1).  ; 

4»  The  older  writers  stated  that  uroleucic  acid  commonly  accompanied  homo- 
gentisic acid  in  the  urine  of  alkaptonuria,  hut  later  observations  do  not  confirm 
this.      (Oswald,  Zeit.  phvsiol.  Chem..  1914    (0.3),  n07 ) . 

■•5  .Tour.  Biol.  Cliem.,  1011    (f)),  130  and  151. 

<«Zeit.  physiol.  Chem.,  1012    (77),  454. 


.\IJ\AI'TOMh'/A  579 

the  appearaiK'c  in  tlio  urine  of  liomogentisic  acid  in  a  normal  individ- 
ual by  feedin<>:  large  amounts  of  tyrosine,  which  is  in  favor  of  the 
view  that  it  is  a  normal  intermediary  in  tj-rosine  catabolism.  In  any 
case  the  administration  of  tyrosine  or  phenylalanine,  or  of  tyrosine- 
rich  foods — i.  e.,  proteins — causes  a  marked  increase  in  the  amount 
of  homogentisic  acid  eliminated  in  the  urine;  indeed,  this  increase  is 
almost  quantitative.  Normal  individuals  when  jjiven  these  substances 
in  moderate  amounts,  or  homogentisie  acid  itself,  destroy  them  com- 
pletely, so  that  the  latter  does  not  appear  at  all  in  the  urine.^"''  If 
alkaptonurics  are  kept  Avithout  protein  food  for  some  time,  the  elimi- 
nation of  alkaptonuric  acids  goes  on,  although  in  diminished  amounts, 
indicating  that  the  aromatic  amino-acids  formed  in  tissue  catabolism 
also  fail  to  be  destroyed  and,  therefore,  appear  in  the  urine  as  these 
derivatives. 

Since  gentisic  acid, 

OH 


COOH, 
HO 

>\'hen  given  by  mouth,  is  also  eliminated  unchanged  by  alkaptonurics, 
although  completely  destroyed  by  normal  individuals,  it  seems  evident 
that  the  difficulty  in  metabolism  affects  the  benzene  ring  itself  and 
does  not  depend  upon  the  character  of  the  side-chain.  Nonnal  organ- 
isms seem  to  be  capable  of  destroying  such  aromatic  compounds  as 
pass  through  a  stage  of  homogentisic  acid  in  being  oxidized,  which 
indicates  that  the  benzene  ring  can  be  broken  up  only  Avhen  oxidized 
in  this  particular  manner  (the  2,  5  position)  ;  the  alkaptonuric  differs 
in  being  unable  to  break  up  even  this  form  (Falta).  According  to 
Garrod  *'  the  conversion  of  tyrosine  and  phenylalanine  into  homogen- 
tisic acid  is  so  complete  that  the  ratio  of  homogentisic  acid  to  nitro- 
gen is  constant  and  the  same  in  all  cases.  Frommherz  and  Her- 
manns ^"'^  advance  the  suggestion  that  normal  oxidation  of  the  aromatic 
radicals  may  take  place  by  two  routes,  one  by  way  of  homogentisic 
acid,  the  other  by  way  of  the  3-4  dioxy-derivatives  (i.  e.,  pyrocate- 
chin),  since  such  derivatives  can  be  readily  oxidized  in  the  metabolism 
of  alkaptonurics  who  cannot  destroy  homogentesic  acid.  That  is, 
their  deficiency  involves  onl}^  one  of  two  possible  methods  of  oxidizing 
aromatic  compounds,  leaving  them  considerable  capacity  for  this  im- 
portant metabolic  function.  The  tissues  of  the  alkaptonuric  are  prob- 
ably not  chemically  affected  in  this  condition,  for  Abderhalden  *^  found 
that  the  hair  and  nails  of  an  alkaptonuric  contained  normal  propor- 
tions of  tyrosine. 

46a  Gross  states  that  normal  scrum  destroys  liomogentisio  acid,  which  property 
is  lacking  in  the  serum  of  alkaptonurics    ( Biochem.  Zeit.,   1914    ( (il ) .   l(i.i). 

47  Garrod  and  Clarke,  T!iocli«>ni.  Zeit.,  1!1()7    (2),  217. 
4"aZeit.   physiol.   Chem..   1914    (01),    194. 

48  Zeit.  physiol.  Chem.,   1907    (52),  435. 


580  GASTRO-INTESTIXAL    "AUTOIXTOXICATIOX" 

In  some  cases  of  alkaptonuria  a  pignuMitation  of  the  cartilages  also 
occurs,  ochronosis,  but  the  association  is  not  constant;  ochronosis  may 
occur  without  alkaptonuria,   and  conversely.     (See   "Ochronosis.") 

(2)    SUBSTANCES  ARISING  FROM  THE  FATTY  ACID   RADICALS 
(AMINO-ACIDS)    OF   PROTEINS 

As  stated  in  the  introductory  chajitei',  the  protein  molecule  con- 
sists of  a  combination  of  a  great  number  of  organic  acids,  of  various 
sorts,  all  of  which  have  as  a  common  characteristic  the  presence  of  an 
NHo  group  attached  to  the  carbon  atom  nearest  the  acid  radical,  the 
a  position;  thus,  R — CHNHo — COOH.  A  few  of  the  amino-acids  con- 
tain an  aromatic  group,  and  the  relation  of  these  to  intestinal  decom- 
position has  been  considered  above.  The  greater  number  have  a 
simple  fatty  acid  radical  (the  simplest  amino-acid  being  glycocoll,. 
CHoNHo — COOH),  and  from  them  are  derived  by  intestinal  putre- 
faction substances  that  are,  for  the  most  part,  chemically  simple  and, 
as  far  as  known,  pathologically  unimportant.  From  leucine  alone  is 
derived  a  substance  of  known  considerable  toxicity,  the  pressor  base 

isoann/linitine. 

>CH  —  CH,  —  CH,  —  NH, 
CH3 

which  is  less  powerful  than  the  cyclic  pressor  bases  described  prev- 
iously.    Bain  '^^^  found  it  the  most  abundant  pressor  base  of  the  urine. 

Fattj^  acids  may  readily  be  formed  from  them  by  splitting  out  of 
the  NH,  group ;  thus  acetic  acid  may  be  formed  from  glycocoll, 
propionic  acid  from  alanine,  etc.  Apparently  butyric  and  acetic  acid 
are  the  acids  most  commonly  formed  in  this  way.  Gaseous  deriva- 
tives, such  as  hydrogen,  ammonia,  carbon  dioxide,  and  marsh-gas,  are 
also  produced.  Acet&ne  is  perhaps  formed  from  these  fatty  acids; 
it  is  often  present  in  the  intestinal  contents,  but  may  come  from  other 
sources. 

Certain  conditions  of  cyanosis  have  been  designated  as  enteroijenous 
cfjanosis,  because  of  tlie  belief  that  the  methem()gh)bin  responsible  for 
the  cyanosis  is  caused  by  nitrites  derived  from  intestinal  putrefaction, 
and  demonstrable  in  the  blood. ^"  Presumably  tlie  nitrites  come  from 
llic  Xll.j  groui)s  of  the  protein  molecule,  the  ('(don  bacillus  being  an 
active  fornuM-  of  nitrites.  Tender  tlie  same  term  are  included  the  cases 
of  sulph-hcinoglohinemia.  This  condition  is  ascribed  by  Wallis"'"  to 
bacteria  which  produce  from  the  proteins  a  hydroxylamine  derivative, 
capable  of  reducing  oxyhemoglobin,  and  whicli  lie  finds  jiresent  in  the 
blood  of  patients  with  sul])li-hemogl()binemia. 

Diamines. — Of  much  interest  are  the  substances  that   arc   formed 

•"■"gwart.   .)()iir.    Kxp.    Plivsiol.,    1014    (S).   2-2!t. 

4(1  Sec    rjilison.    Quart.    Jour.    :Mo(1.,    100"     (1).    -i!) :    Wost   and    Clarke.    Laiieet, 
Feb.  2,  in(»7;   Davis.  Lancet.  Oct.  2ti.  VM'l. 
•-■•o  Quart.   Jour.    Med.,  Oct.,    ]!»!:{. 


i>i:in\  .\Ti\  i:s  of  siu'iiiu  rosTAisisa  I'ifOTEiN  radicals       581 

from  tlio  ainino-aeids  by  bacterial  action,  wliich  still  retain  their  nitro- 
gen radicals — the  ptoindins.  Two  of  these,  the  diamines  putrescine, 
NII2  (CIIn)4  NIL,  and  cadaverine,  Nil,  (0X12)5  NIIj  are  of  particular 
interest,^^  because  they  have  been  observed  in  the  feces  and  urine  of 
persons  with  cystinurw.  The  stools  in  cholera  also  seem  to  contain 
these  ptoma'i'ns  frequently.  Their  etiological  relation  to  the  cystinuria 
is  no  longer  accepted,  however,  and  their  toxicity  is  slight.  They  are 
probably  derived  from  the  diamino-acids  of  the  protein  molecule,  pu- 
trescine being  closelj'  related  to  ornithine,''^''  and  is  probably  formed 
from  it  as  follows  : 

NHa  NHj  NHo  XH, 

CH2  —  CH,  —  CH,  —  CH  —  COOH  p=,  CH,  —  CTI,  —  CH,  —  CH,  +  CO, 
( ornithine )  (  putrescine ) 

while  cadaverine  is  probably  formed  from,  lysine, 

NH„  NHo  NH2  NH, 

CH,—  (CH,),  — CH  — COOH  .=.  CH,—  (CH,),  — CH,  +  CO, 
(lysine)  (cadaverine) 

.NH, 
Ethylidendiamine,  CH^-CH  <  which  is  somewhat  toxic,  has  also 

^NH,, 
been  detected  in  the  contents  of  the  gastro-intestinal  tract. 

Apparently  these  substances  are  absent  from  normal  feces,  but 
this  does  not  exclude  the  possibility  of  their  normal  formation,  ab- 
sorption, and  destnietion.  There  is  no  evidence  that  they  ever  cause 
symptoms  or  pathological  alterations. 

(3)      SUBSTANCES    ARISING    FROM    THE    SULPHUR-CONTAINING    RADICAL 

OF  PROTEINS 

Most  if  not  all  of  the  sulphur  in  the  protein  molecule  seems  to  be 
contained  in  the  amino-acid.  cystine,  which  has  the  following  compo- 
sition : 

S  —  CH,  —  CHXH,  —  COOH 

S  —  CH,  —  CHNH,  —  COOH. 

From  this  is  formed  the  hydrogen  sulphide  of  the  intestinal  gases,  of 
which  about  0.058-0.066  gram  is  present  in  each  one  hundred  grams 
of  normal  colon  contents.  Although  Senator  has  described  a  case  in 
which  an  intoxication  with  HoS  of  intestinal  origin  occurred,  this 
gas  seems  not  to  be  a  frequent  cause  of  intoxication,  and  Senator's 
case  stands  almost  alone.  Under  normal  conditions  H^S  does  not 
appear  in  the  urine,  any  that  may  be  absorbed  probabh^  being  oxidized 

51  For  discussion  of  formation  and  properties  of  these  t\vo  ptomains,  see 
Vanghan  and  No\'\''s  ''Cellular  Toxins." 

51a  Ornithine  forms  part  of  tlie  arginine  molecule,  which  is  the  most  universally 
present  (in  proteins)  of  all  tlie  amino-acids,  ornithine  being  formed  when  urea 
is  split  from  arginine. 


582  GASTRO-INTESTIKAL    'AUTOIXTOXICATIOX" 

to  SO4.  If  enough  H2S  should  enter  the  blood  so  that  it  was  not 
completely  destroyed,  it  might  well  cause  harm,  for  it  is  decidedly 
toxic,  particularly  att'ecting  the  nervous  system;  but  we  have  no  evi- 
dence that  this  often  happens.  Van  der  Bergh  ^-  has  observed  cases 
of  intestinal  obstruction  in  which  the  presence  of  sulphemoglo'bin  in 
the  patient's  blood  was  demonstrated. 

Methyl  mercaptan,  CII3SII,  has  also  been  found  in  the  feces,  al- 
though it  seems  not  to  be  abundantly  or  constantly  present,  according 
to  Herter,^^  who  found  also  that  mixed  bacteria  from  normal  feces 
rarely  produce  mercaptan  in  cultures.  However,  bacteria  from  the 
feces  of  persons  suffering  with  various  diseases  often  produce  mercap- 
tan. Ethyl  mercaptan,  CoH-SH,  and  ethyl  sulphide,  CoHj-S-CjHj, 
have  also  been  described  as  fecal  constituents.  It  is  not  known  that 
the  mercaptans  are  a  cause  of  intoxication. 

CYSTINE  AND  CYSTINURIA  '■* 

The  presence  of  cystine  in  the  urine  has  been  observed  in  a  num- 
ber of  cases,  and  when  present  at  all  it  is  usually  present  in  consider- 
able quantities.  Because  of  its  slight  solubility  it  appears  as  a  de- 
posit of  hexagonal  crj'stals,  and  frequently  forms  cystine  concretions 
(q.  V.)  in  the  urinary  bladder."'^  According  to  Garrod  it  is  more 
common  than  alkaptonuria,  and,  like  the  rest  of  the  "Inborn  Errors 
of  Metabolism,"  occurs  much  more  often  in  males  than  in  females. 
Hofmann  ^^  was  able  to  collect  from  the  literature  to  1907  a  total  of 
175  cases,  of  which  85  were  males  and  45  females.  Baumann  and 
others  observed  that  in  cystinuria  the  urine  often  contains,  besides 
the  cystine,  the  diamines  cadaverine  and  putrescine,  which  are  formed 
from  lysine  and  ornithine  respectively  in  the  intestines  througli  putre- 
faction, and  they  naturally  suspected  that  cystine  arose  in  the  same 
way.  Another  view  was  that  the  diamines  interfered  with  the  oxida- 
tion of  sulphur  in  the  body,  so  that  it  was  eliminated  in  the  unoxidized 
form  of  cystine.  But  it  has  been  demonstrated  that  neither  of  these 
hypotheses  is  correct,  for  (1)  cystine  could  not  be  found  in  the  feces; 
(2)  if  given  by  mouth,  it  is  completely  oxidized,  and  causes  only  the 
appearance  of  excessive  amounts  of  sulphates  in  the  urine;  (3)  cys- 
tinuria has  been  observed  to  occur  independent  of  the  presence  of 
the  diamines,  and  not  to  be  modified  or  caused  by  their  administration 
or  pathological  formation.  The  view  now  prevalent  is  that  the  cystine 
that  escapes  in  the  urine  in  c^-stinuria  is  not  derived  from  intestinal 

52Deut.  Arch.  klin.  Med.,  1905    (S3),  SG. 

53  .Tour.  Biol.  Cliem.,  1906   (1),  421. 

54  Literature  coneerninfj  cystine  {,'ivoii  l)y  Fricdmanii,  Kifiohiiissc  der  Physiol., 
1902  (I.  Aht.  1).  If);  and  l)y  Maiui.  "Chemistry  of  the  Proteins."  jjp.  r)()-(i4.  Cys- 
tinuria reviewed  l)y  I5(')dtker.  Zeit.  ])hysiol.  Chom.,  1905  (45),  393;  C.arrod,  "Inborn 
Errors  of  Metaholism,"  and  Lancet,  duly,  P.tOS. 

5-'>  Ahderhaldcn    (Zeit.   physiol.   Chem.,    1003    I3S).  f).!?)    has  described  a   ease   in 
a  child  in  which  tlie  or^ians  were  inliJtrated  witii  masses  of  tlie  cystine  crystals. 
ooCent.  Grenz.  Med.  u.  Chir.,  1907    (KD.  721. 


i'i,-(U)i  <'rs  or  Fh'h'\ii:\TAT/<>.\  or  <•  AUiioiiyiu; M-rn"        583 

putrefac'tioiu  l»ut  is  formed  in  the  tissues  from  the  protein  molecule, 
and  fails  to  he  further  decomposed  because  of  some  anomaly  of  metab- 
olism. This  view  is  supported  by  the  fact  that  cj'stinuria  often  ap- 
pears to  be  an  hereditary  disease,  occurrino:  in  families  for  several 
jivnerations;  it  is  independent  of  the  diet,  cystine  appearin<>-  even  if 
proteins  are  withheld,  and  also  independent  of  intestinal  putrefac- 
tion."'" It  having  been  found  that  leucine  and  tyrosine  may  also  occur 
in  the  urine  in  cystinuria,  it  seems  probable  that  this  condition  de- 
pends upon  a  general  abnormality  of  protein  metabolism.  The  rela- 
tion of  the  diamines  to  the  condition  is,  however,  very  uncertain. 
Cystine  does  not  seem  to  exert  any  toxic  effect,  and  patients  with 
cystinuria  do  not  usually  appear  to  sutfer  greatly  from  the  abnormal 
metabolism,  the  chief  trouble  observed  being  due  to  the  formation  of 
the  concretions  in  the  bladder.""''  Sometimes  in  children,  however, 
emaciation  and  early  death,  without  other  apparent  cause,  have  been 
observed,  and  may  be  due  to  the  metabolic  anomaly. 

The  metabolic  error  in  cystinuria  is  not  complete,  for  only  a  por- 
tion of  the  total  cystine  of  the  catabolized  proteins  is  excreted  as 
such  (Garrod).  This  would  amount  to  some  five  grams  per  day, 
whereas  the  average  excretion  is  only  about  0.3-0.5  gram,  and  sul- 
phates and  other  neutral  sulphur  compounds  are  always  present  in 
the  urine.  In  no  condition  other  than  cystinuria  have  putrescine  and 
cadaverine  been  found  in  quantities  which  could  be  detected  by  ordi- 
nary methods  in  2-4-hour  specimens ;  they  may  also  be  found  in  the 
feces  of  cystinurics,  where  cystine  is  never  found.  In  the  urine  their 
presence  is  inconstant,  and  the  amounts  are  at  best  very  small.  Leu- 
cine and  tyrosine  are  found  much  less  often  than  the  diamines ;  lysine, 
has  been  found  in  one  case,^*  which  supports  the  view  that  cadaverine 
and  putrescine  come  from  the  diamino-acids  of  the  protein  molecule 
by  metabolism  rather  than  by  putrefaction. 

B.     PRODUCTS  OF  FERMENTATION  OF  CARBOHYDRATES 

These  include  practically  all  the  members  of  the  fatty  acid  series, 
from  formic  acid  to  valerianic  ackl;  and  the  oxy-acids,  lactic,  succinic, 
and  o.ryhutijric;  also,  oxalic  acid,  acetone,  ethyl  alcohol,  and  the  fol- 
lowing gases:  CO^,  CH^,  H,.  For  the  most  part,  the  various  organic 
acids  are  absorbed  through  the  intestinal  walls,  and  are  oxidized 
completely  in  the  tissues  without  causing  any  harm  whatever.  The 
possibility  that  acid  intoxication  may  be  produced  in  this  way  has 
been  suggested,  but  it  is  generally  believed  that  this  does  not  occur, 
except  possibly  in  infants.     Lactic  and  butyric  ''^^  acids  are  formed 

57  An  isolated  case  of  transient  cystinuria  in  a  patient  with  Raynand's  disease 
is  described  by  Githens   (Penn.  ^led.  Jour.,  1910   (1),  507). 

57a  This  may  be  avoidetl  by  decreasinrr  the  cystine  by  means  of  a  low  jirotein 
diet,  and  increasinof  its  solubility  by  keeping  the  reaction  of  tlie  urine  alkaline 
(Smillie.  Arch.  Int.  Med..  1915    (IG),  50.3). 

•''•'^  Ackermann  and  Kutscher,  Zeit.  f.  Biol.,   1011    (57),  355. 

5sa  Coleman    (Ann.    Inst.    Pasteur,    1915    (29),    139)    attempts   to   incriminate 


584  GASTh'oi\ri:sTJ.\AL  '-AUToiy toxic ATioy 

particularly  in  gastric  fermentations  in  persons  with  deticient  hydro- 
chloric acid,  motor  insufficiency,  or  or<>-anic  obstruction.  ]\Iost  of  the 
disturbances  observed  in  these  conditions  seem  to  be  due  to  distention 
of  the  stomach  with  gas,  chiefly  CO2,  which  is  formed  during  the  fer- 
mentation. It  is  possible,  however,  that  the  organic  acids  exercise 
some  irritant  effects  on  the  mucous  membrane;  and  they  may  also 
cause  diarrhea,  lactic  and  acetic  acid  often  being  present  in  diar- 
riieal  discharges  due  to  excessive  feeding  with  carbohydrates 
(Herter). 

These  acids  or  their  salts  do  not  appear  in  the  urine,  unless  possibly 
as  minute  traces,  except  the  oxalic  acid.  ^linute  quantities  (0.02  gm. 
per  day)  of  this  substance  are  present  in  normal  urine,  but  larger 
quantities  (oxaluria)  seem  to  depend  either  upon  the  taking  of  food 
containing  much  oxalic  acid  (rhubarb,  spinach,  etc.)  or  upon  excessive 
gastric  fermentation  of  carbohydrates  (Baldwin),"''  and  perhaps  upon 
excessive  destruction  of  purines,  from  which  oxalic  acid  may  be  de- 
rived. .  Of  the  amino-acids  it  is  presumably  the  diatomic  acids,  glu- 
tamic and  aspartic,  which  yield  oxalic  acid  (Jastrowitz).*'^*  Others, 
however,  do  not  admit  that  any  appreciable  amount  of  oxalic  acid  is 
derived  from  proteins.'^"-''  Probably  the  small  quantities  of  oxalie 
acid  thus  formed  do  not  cause  toxic  etfects,  and  are  important  chiefly 
as  causing  urinary  concretions  of  calcium  oxalate,  although  there  is. 
evidence  that  long-continued  excretion  of  oxalic  acid  may  cause  renal 
lesions.      (See  also  consideration  of  oxalic  calculi.  Chap,  xv.) 

C.     PRODUCTS   OF  THE   DECOMPOSITION  OF  FATS 

These  differ  but  little  in  nature  from  the  products  of  carbohydrate 
fennentation,  the  large  fatty  acid  molecules  being  broken  down  to 
smaller  ones.  In  infants  these  fatty  acids  have  been  believed  to  be  a 
cause  of  acid  intoxication  and  acetonuria,"^  but  probably  they  are  sel- 
dom, if  ever,  of  pathological  importance.  It  is  possible,  however,  that 
a  serious  reduction  in  the  bases  of  the  blood  may  result  from  the 
formation  of  excessive  amounts  of  fatty  acids  in  the  intestines,  the 
bases  being  combined  to  unite  with  the  fatty  acids,  and  then  excreted 
in  the  feces. 

It  is  quite  otherwise  with  the  products  of  decom])ositi()n  of  leci- 
thin.^^     From  its  molecule  is  split  o1"f  the  ptonui'in,  cJwlinc, 

{(  11:,) ,.  =  X  —  C'll.  —  CH,OH, 

I 
OH 

Imtyric  acid  in  tlic  production  of  iutorioscloiosis,  while  Oswald  Loeh  believed 
lax-tic  at'id  to  be  important,  a  "  'e-v  wliicli  could  not  be  alto<retiier  supjiorted  by 
Denny  and  Frotliinj,diam,  .lour    Mel.  Kes.,   I!tl4    (.'?1).  277. 

■'•".Tour.   Exp.   Med.,   IHOO    (.')),  27. 

"0  ]?ioclieni.  Zeit.,   1!)10    (28),  ;U. 

ooa  \Ve;xrzyno\vski,  Zeit.  pliysiol.  Clieni..  lOl.S   (  S.'i ) .  112. 

"1  Meyer  and  l^anjistein,  Jalirb.  f.  KinderlieilU.,   \'MH\    id:!).  ^0. 

"-Literature  given  by  Halliburton,  Kr^ebniisse  der  I'liysiol..   l!M)4    (4).  24. 


RESULTS  or  (;.\sTi,'<)-i\Ti:sr/\\/.  imoxkatios  585 

which  is  easily  oxidizt'd  into  a  lii^^lily  poisonous  eoiiipoiiiid,  isomeriu 
with  luuscariite,  of  hy  h)sin<i-  a  mok'cule  of  water  it  forms  ncurine, 

(C'll3),=  X  — ('11  -  L'U,, 

Oil 

which  is  also  very  poisonous  (discussed  under  ^'Ptomai'ns, "  Chap.  iv.). 
It  has  been  demonstrated  by  Nesbitt ""  that  in  the  contents  of  ob- 
structed intestines  of  dogs  that  have  been  fed  lecithin-rich  food  (eggs) 
both  choline  and  neurine  may  be  found  free,  and  Kutscher  and  Loh- 
mann  "*  have  found  neurine  in  human  urine.  It  seems  possible  that 
some  of  the  toxic  effects  observed  after  eating  excessively  of  such 
food. as  calves'  brains,  or  eggs,  may  depend  upon  intoxication  with 
the  products  of  lecithin  decomposition.  Also,  the  normal  presence 
of  trimethj^lamine  in  the  blood  and  cerebrospinal  fluid  (Doree  and 
Golla)"''  may  be  from  this  source.  Hunt,''^"''  who  has  done  extensive 
Avork  with  choline,  states  that  at  present  we  have  no  grounds  for 
believing  that  choline  has  any  significance  in  physiological  or  patho- 
logical processes.  There  is  no  evidence  that  the  highly  active  acetyl- 
choline •'^''  is  produced  from  choline  in  the  body,  but  in  view  of  the 
enormous  toxicity  of  this  choline  derivative  there  must  always  be  con- 
sidered the  possibility  that  such  toxic  choline  compounds  may  at  times 
develop  in  amounts  too  small  to  be  detected  but  large  enough  to  cause 
severe  effects. 

RESULTS  OF  GASTRO-INTESTINAL  INTOXICATION 

As  we  have  seen  from  the  above,  but  few  of  the  known  products  of 
gastro-intestinal  putrefaction  are  toxic  to  any  considerable  degree, 
and  these  are  probably  produced  in  too  small  quantities  to  cause  any 
appreciable  effect,  especially  in  view  of  the  detoxicating  and  elimi- 
natory  powers  of  the  liver,  kidney,  and  other  organs.  And  yet  we 
have  abundant  clinical  evidence  that  excessive  intestinal  putrefaction 
or  retention  of  the  intestinal  contents  causes  marked  disturbance  in 
health.  The  slight  malaise,  headache,  and  lassitude  observed  as  the 
result  of  simple  constipation  may  possibly  be  adequately  accounted  for 
by  intoxication  with  indole  and  similar  substances,  although  we  have 
no  conclusive  proof  that  such  is  the  case.  Two  explanations  may  be 
suggested :  One  is  that  the  intestinal  flora  becomes  altered  because  of 
the  changed  conditions,  and  bacteria  thrive  that  produce  specific 
soluble  toxic  substances,  analogous  to  those  formed  by  B.  botulinus, 
or  similar  to  the  tyrotoxicon  (Vaughan)  that  may  be  formed  in  milk 

63  Jour.   Exp.   Mod.,    1S99    (4),    1;    see  also  Iloesslin,   Hofmeister's   Beitr.,    190(> 
(8),  27. 
o-iZeit.  physiol.   Chem.,   1906    (48),   1. 
6-.  Biochem.  Jour..   1910    (5),  306. 
65a  Jour.   Pharmacol..    191;)    (7),  nOl. 
65b  See  Dale,  Jour.  Pharmaeol.,  1914   (6),  147. 


586  (V .  1 N Th'o-ix Ti:^ ri vi /,  '•. i ltoix toxic. i tiox" 

and  luilk  products.  Thus  Clairinoiit  and  Raiizi ""  found  lieat-resistant 
toxic  substances  in  the  intestinal  contents  in  ileus  (experimental),  and 
similar  substances  could  also  be  obtained  b}'  growing  cultures  of  the 
intestinal  contents  on  bouillon.  Another  explanation  is  that  many 
unidentified  poisonous  substances  are  produced  in  the  alimentary 
canal  which  ordinarily  are  destroyed,  but  under  certain  conditions 
may  be  reabsorbed.  That  unrecognized  toxic  substances  are  formed 
in  the  intestines  is  almost  certain,  for  it  has  been  repeatedly  shown 
that  extracts  of  the  contents  of  the  alimentary  canal  are  very  poison- 
ous. Although  the  teehnic  of  many  of  these  experiments  has  been 
questionable,  the  results  have  been  obtained  so  often  as  to  render  it 
probable  that  the  main  contention  is  correct."^'  Thus  ^lagnus-Alsle- 
ben '^"'^  has  found  in  the  upper  part  of  the  small  intestine  of  dogs  (ex- 
cept when  on  milk  diet)  a  very  poisonous  substance  which  killed  rab- 
bits by  respiratory  paralysis,  but  which  is  inert  when  injected  into  the 
portal  vein.  Extracts  of  the  wall  of  the  large  intestine  are  also  toxic, 
and  lose  their  toxicity  at  60°,  by  passing  through  porcelain  filters  and 
by  treatment  with  alcohol ;  extracts  of  fetal  intestines  are  not  toxic 
(Distaso)."^ 

Whipple  ""  has  demonstrated  that  closed  duodenal  loops  in  dogs 
come  to  contain  a  highly  toxic  substance  of  unknown  nature,  appar- 
ently formed  in  the  epithelium  of  the  gut  rather  than  in  its  contents, 
which  causes  severe  splanchnic  congestion,  vomiting  and  diarrhoea 
when  injected  into  normal  dogs.  The  agent  is  not  destroyed  hy 
autolysis,  filtration  or  heating  at  60°,  yet  dogs  can  be  made  somewhat 
immune.  The  origin  and  nature  of  this  poison  have  not  yet  been  de- 
termined, but  it  seems  probable  that  it  is  an  important  factor  in  the 
intoxication  of  intestinal  obstruction.  Apparently  the  liver  does  not 
have  much  detoxicating  ett'ect,  for  dogs  with  p]ck  fistula  behave  much 
the  same  when  the  intestine  is  obstructed  as  normal  dogs.  A  similar 
material  cannot  be  obtained  by  hydrolysis  or  autolysis  of  normal  duo- 
denal mucosa,  the  obstruction  being  an  essential  feature.  Obstruction 
of  lower  portions  of  the  intestine  has  much  less  effect,'^  and  it  has 
been  suggested  that  the  poiscm  formed  in  the  duodenum  is  neutralized 
or  destroyed  farther  down  in  the  intestine.^- 

In  any  case,  correctly  or  incorrectly,  a  great  num])er  of  disease  con- 
ditions have  been  attributed  to  poisons  of  gastro-intestinal  orig'in, 
including  not  only  such  minor  conditions  as  headache,  nuilaise,  lassi- 
tude, etc.,  but  also  sciatica,  tetany,  epilepsy,  eclampsia,  many  forms 
of  dermatitis,  various  forms  of  nervous  diseases,  myxedema  and  cretin- 

"«  Arch.  klin.  Chir.,  1904    (73),  090. 

(17  For  example,  see  Roijer  and  (ianiicr,  Coiiipt.  I\eiul.  Soc.  IJiol.,  190.")  {'i9) , 
388  and  f)74 ;   190(1   (60),  CM). 

OS  IlofmeiBter's   Beitr.,    190.5    (fi),   i)0.'l. 

If  Zcit.  Inimunitiit.,   191:5    (10),  400. 

■'I  \\liil)j)le.  Sloiie  and   Bcrnlieini.  .lour.   K\]H'r.  ^\^H\..   19l;i    (17),  280. 

71  See   liimtiii^'.  .(..iir.   K\]u-v.   Mod.,    1 9 1  ;i    (17).    192. 

72  Maury,   .\iiici-.  .Idiii-.   Med.  S.-i.,    19i)<l    (l;i7l,  72'). 


RESULTS  OF  (!.l,STh'<>-l\Ti:sT/\\l.   I  \T()\  l<!.\TI()S  587 

ism,  clilorosis  aiul  pernicious  auciiiia,  cirrhosis,  ii('])lii-itis,  and  arterio- 
sclerosis.'" Wliile  in  many  cases  the  severity  of  tliese  various  eou- 
ditioiis  is  apparently  augmented  by  intestinal  disturbances,  the  etio- 
logic  relation  is  not  so  clear.  That  long-continued  intoxication  of  in- 
testinal origin  may  cause  serious  injury  to  the  tissues  is,  however, 
extremely  probable.  There  is  much  reason  for  believing  that  many 
cases  of  non-alcoholic  cirrhosis  are  due  to  this  cause;  not  improbably 
chronic  nephritis,  myocarditis,  and  arteriosclerosis  may  occasionally 
be  the  result  of  long-continued  intoxication  from  the  same  source. 
Arteriosclerosis  especially  has  been  attributed  to  indole  and  related 
substances  by  ^letchnikoft'  and  his  associates,  who  have  produced  ar- 
teriosclerosis in  rabbits  by  injecting  indole,  but  not  with  skatole.  As 
is  well  known,  ^letchuikoft*  believed  that  most  of  the  manifestations  of 
senility  come  from  putrefaction  in  the  large  bowel,"*  and  a  number 
of  observers  have  described  as  products  of  intestinal  putrefaction  cer- 
tain pressor  substances  of  high  potency  which,  presumably,  might 
cause  serious  arterial  and  cardiac  injury.'^  An  elaborate  study  of 
a  certain  type  of  cases  of  defective  development  led  Herter  '•*  to  the 
conclusion  that  intestinal  intoxication  is  responsible,  and  hence  he 
designated  this  condition  "intestinal  infantilism." 

Tetany  associated  with  gastric  dilatation  offers  perhaps  the  strongest 
case,  numerous  observers  having  reported  finding  a  marked  toxicity 
of  the  stomach  contents.'^^  Pineles  ^^  considers  that  all  forms  of  tet- 
any, whether  of  gastric  origin  or  following  thyroidectomy,  are  due 
to  one  and  the  same  "tetany  poison." 

Although  there  are  usually  evidences  of  intoxication  in  acute  dila- 
tation of  the  stomach,  yet  there  is  no  good  evidence  as  to  its  nature. 
It  is  suggested  by  Woodyatt  and  Graham  "^  that  the  dilatation  is  pro- 
duced by  CO,  secreted  into  the  stomach  from  its  walls.  There  is  also 
considerable  evidence  that  the  tetany,  when  present,  is  associated 
with  a  deficiency  in  calcium  in  the  blood  and  nervous  tissue,  and 
that  this  is  further  related  to  the  functional  activity  of  the  parathy- 
roids {q.  I'.). 

The  relation  of  intestinal  intoxication  to  the  various  anemias,  par- 
ticularly chlorosis  and  pernicious  anemia,  has  been  repeatedly  indi- 
cated and  discussed.  Clinical  evidence  strongly  indicates  that  such  a 
relation  exists,  and  there  is  no  doubt  that  hemolytic  substances  may 
be  formed  in  the  alimentary  tract, ^°  but  that  chlorosis  and  pernicious 

T3  The  relation  of  orastro-intestinal  intoxicatinn  to  these  various  diseases  is  re- 
viewed by  Weintraud,  Ergeh.  alio:.  Pathol.,  IS!)?    (4),   17. 

74  See  Ann.  Inst.  Pasteur,  1010    (24).  75.i. 

75  See  Granger,  Arch.  Int.  :\red.,   1!)12    (10).  202. 

Tfi  See  McCnidden,  .Jour.  Exper.  INled.,  1012   (If)),  107. 

"Bibliography  bv  Halliburton  and  McKendrick.  P.rit.  M.d.  -Tour.,  inoi  (i), 
1G07. 

78Deut.  Arch.  klin.  Med.,   1006    (S.5).  401. 

79  Trans.  Chicago  Path.  Soc.   1912    (8),  3.")4. 

80  See  Kiilbs,  Arch,  exper.  Path..  1006  (o.5),  7.3:  also  Herter,  .Tour.  P.iol.  Cheni.. 
1906   (2),  1. 


588  GASTRO-INTESTIXAL    "AUTOiyTOXICATIOX" 

anemia  do  depend  upon  intestinal  putrefaction  or  infection  is  far 
from  established   (see  ''Anemia."  Chap.  xi). 

It  seems  highly  probable  that  o-astro-intestinal  "autointoxication" 
would  be  a  much  more  serious  matter  were  it  not  for  the  mechanisms 
of  defence  possessed  by  the  body,  especially  in  the  liver.*^  For  exam- 
ple, Richards  and  Howland  have'  indicated  the  increased  toxicity-  of 
indole  when  the  oxidizing  ])ower  of  the  liver  is  reduced,  and  Herter 
and  Wakeman  have  sliown  the  power  of  the  liver  to  combine  indole 
and  thus  remove  it  from  circulation.  This  topic  has  been  discussed 
more  fully  elsewhere  (Chap.  ix). 

ACUTE  INTESTINAL  OBSTRUCTION 

The  violent  effects  that  follow  complete  occlusion  of  the  intestine, 
especially  in  the  upper  portion,  must  be  due  to  some  highly  toxic  sub- 
stance or  substances.  The  clinical  features  of  obstructive  ileus, 
namel}',  vomiting,  collapse,  complete  muscular  relaxation,  and  sub- 
normal temperature,  are  associated  with  the  excretion  of  larg-e  quan- 
tities of  indican  and  other  substances  combined  with  sulphuric  acid, 
proving  that  intestinal  putrefaction  is  active.  Undoubtedly  in  ileus 
we  have  a  profound  and  rapidly  fatal  intoxication  with  substances 
formed  in  the  obstructed  intestines. 

Whipple  '"  has  demonstrated  that  closed  duodenal  loops  in  dogs 
come  to  contain  a  highly  toxic  substance  of  unknown  nature,  appar- 
ently formed  in  the  epithelium  of  the  gut  rather  than  in  its  contents, 
which  causes  severe  splanchnic  congestion,  vomiting  and  diarrhoea 
when  injected  into  normal  dogs.  The  toxic  ag'ent  is  not  destroyed  by 
autolysis,  filtration  or  heating  at  60°,  yet  dogs  can  be  made  somewhat 
refractory  or  immune.  From  the  contents  of  such  loops,  and  from  the 
bowel  above  obstructions,  he  has  isolated  a  very  toxic  proteose,'*-  which 
he  believes  may  be  responsible  for  the  intoxication.  Wliether  this 
proteose,  or  Avhatever  the  active  poison  may  be,  comes  from  bacterial 
infection,  autolysis,  duodenal  secretion,  or  what,  is  not  yet  agreed  by 
the  numerous  investigators  in  this  field.-^  Apparently  the  liver  does 
not  have  much  detoxicating  effect,  for  dogs  with  Eck  fistula  behave 
much  the  same  when  the  intestine  is  obstructed,  as  dogs  with  normal 
circulation.  A  similar  material  cannot  be  obtained  by  hydrolysis  or 
autolysis  of  normal  duodenal  mucosa,  tiie  obstruction  being  an  essen- 
tial feature.  Obstruction  of  lower  portions  of  the  intestine  has  much 
less  effect,^^  and  it  has  been  suggested  that  the  poison  formed  in  the 
duodenum  is  neutralized  or  destroyed  farther  down  in  the  intestine.^- 
A  striking  feature  of  intestinal  obstruction  is  the  high  non-protein 

*<i  For  disc'ussiiin  and  literature  see  Lust.  Tlofmeister's  Beitr.,  100,5    (G),  1,32. 

■^-'.Tour.  Amer.  Med.  .\ss(m-..  1!)15  (1)5),  470:  19Hi  ((>7),  IT);  Jour.  Exp.  Med., 
1910   (23),  12:{:   l!tl7   (2.")),  2:51  and  401. 

«:<  See  Murpliy  and  IJrooks.  Areli.  Int.  Med..  ini.->  (l.")),  .302;  Curd..  .Tour. 
Infect.  Dis..  1014  (1.')).  124:  Draper.  .Tour.  Anier.  Med.  Assoe..  1010  (67),  1079; 
Dragstadt,  Moorliead  and  Hurcky.  .lour.  K\p.  ^led..  1017   (2.1),  421. 


ACUTE  INTESTINAL  OBSTRUCTIOS 


589 


nitrog-en  content  uf  tlu-  l.lo.ul,  W^nvv.  similar  to  those  of  fatal  uremic 
coma  being  common-  which  may  be  the  result  ot  absor-ption  of  c  eav- 
age  products  from  the  intestine,  or  toxogenic  destruction  ot  tissue 
proteins,  or  both. 

S4  Cooke,  Rodenbaugl.  and  Whipple,  Jour.  Exp.  Med.,  191G   (23),  123. 


CHAPTER   XX 

CHEMICAL  PATHOLOGY  OF  THE  DUCTLESS 

GLANDS  1 

DISEASES  OF  THE  THYROID  - 

As  we  liave  miieh  evidence  that  the  thyroid  has  a  marked  influence 
upon  metabolism,  and  also  that  it  may  be  of  importance  in  preventing 
autointoxication,  the  chemistry  of  diseases  of  the  thyroid  may  be  ap- 
propriately considered  in  connection  with  the  autointoxications. 

THE  FUNCTIONS  OF  THE  THYROID 

Metabolic  Function. — That  the  thyroid  has  an  important  relation 
to  metabolism,  especially  of  proteins,  is  shown  by  the  following-  facts : 

(1)  Administration  of  the  gland  substance,  or  active  preparations 
made  from  it,  to  healthy  men  or  animals,  causes  a  greatly  increased 
elimination  of  nitrogen  in  the  form  of  urea.  This  nitrogen  comes  not 
only  from  the  food,  but  also  from  increased  tissue-destruction,  as  is 
shown  by  the  loss  of  weight  and  strength,  and  by  the  increased  ex- 
cretion of  sulphur  and  phosphorus.  An  increased  destruction  of  the 
body  fat  also  occurs,  so  that  thyroid  therapy  has  been  found  efficient 
in  the  treatment  of  obesity,  but  often  dangerous  because  of  the  rela- 
tively great  amount  of  tissue-destruction.  Basal  metabolism  is  most 
markedly  raised  in  hyperthyroidism,  and  is  lower  in  cretinism  and 
myxedema  than  in  any  other  disease.-'' 

(2)  Loss  of  thyroid  tissue,  either  through  operation  or  disease, 
greatly  reduces  both  nitrogenous  metabolism  and  oxidative  processes. 
Administration  of  thyroid  preparations  under  these  conditions  will 
bring  the  nitrogen  elimination  and  the  gas  exchange  back  to  normal. 

(3)  Deficient  thyroid  secretion  in  young  animals  prevents  their  de- 
veloping normally,  the  amount  of  deficiency  varying  from  nearly  total 
lack  of  development  in  extreme  cretinism  to  slight  grades  of  defective 
development  (infantilism)  or  delayed  maturity.  In  adult  animals, 
besides  decreased  metabolism  there  occur  also  various  trophic  changes 
in  the  skin  and  its  appendages,  an  increased  amount  of  mucin-like 
material  in  the  tissues,  and  greatly  decreased  nervous  and  mental  ac- 

1  Thorough  reviews  of  tin-  ciitiic  sulijcct  (if  llic  diictlfss  glands  nro  lmvcii  hy 
Biedl,  "Tnnero  Sekretioii,"'  rrhaii  and  Scliwar/ciilK'rjr.  r>crlin.  ini;i:  and  Vincent, 
Erffchnissc  I'livsioi..   lltlo   (!)),  451;    11)11    (10).  21S. 

-  Coiicerninfr  tlie  thyroid  see  besides  iiiedl  and  \'ineent,  the  review  bv  llirelier, 
Ergebnisse  Pathol.,   1011,  XV    (,),  82. 

iiaDu  Bois,  Arch.  Int.  Med.,  1916    (17).  i)15. 

590 


77//;  Fr\(Tio\s  or  Tin-:  Tiirnoih  591 

tivity.  All  these  conditions  are  relieved  to  jrreater  or  less  degree  by 
adiniiiistratioii  of  thyroid  tissue  or  its  preparations.'*  Evidently, 
therefore,  tiie  thyroid  exerts  an  influence  upon  growth  and  tissue 
changes ;  whether  this  depends  upon  its  influence  upon  metabolism,  or 
is  an  independent  and  specific  function,  cannot  be  determined.* 

IIow  the  thyroid  or  its  secretion  modifies  metabolism  is  not  yet  un- 
derstood. One  is  reminded  of  the  effects  of  kinases  upon  enz^-mes 
and  their  antecedents,  and  it  may  be  imagined  that  the  thyroid  secre- 
tion activates  both  proteolytic  and  oxidative  enzymes  vv^ithin  the  cells. 
Shryver,"^  indeed,  did  find  that  administration  of  thyroid  to  dogs  for 
some  time  before  killing  them  causes  their  liver  tissue  to  undergo 
autolysis  more  rapidly  than  normal,  although  AVells "  had  been  unable 
to  observe  any  increased  amount  of  autolysis  when  thyroid  extracts 
acted  upon  liver  tissue  in  vitro.  Experimental  observations  show 
that  carbohydrate  metabolism  is  much  influenced  by  the  thyroid,  so 
that  thyroidectomized  animals  may  fail  to  show  glyeosviria  from  vari- 
ous procedures  that  usually  produce  it  (King),^  and  they  are  incapable 
of  utilizing  sugar  injected  parenterally  as  well  as  normal  animals ;  ^ 
they  also  exhibit  an  excessive  creatine  output,  but  otherwise  show  no 
striking  changes.'*'' 

Detoxicatory  Function. — The  evidence  that  the  thyroid  has  for  its 
function  the  destruction  or  neutralization  of  poisonous  substances 
formed  in  metabolism  or  through  intestinal  putrefaction  is  as  follows : 

(1)  After  total  removal  of  the  thyroid  from  many  species  of  ani- 
mals acute  symptoms  develop  that  suggest  strongh'  an  intoxication. 

(2)  After  removal  of  the  thyroid,  marked  changes  occur  in  the 
blood,  there  being  a  severe  anemia  (as  low  as  2,000,000  red  corpus- 
cles), Avith  some  leucocytosis,  and  there  occur  structural  changes  in 
the  blood-vessel  walls  (Kishi).'*  Cytoplasmic  degeneration  of  the 
liver,  kidneys,  and  myocardium  may  also  result  (Bensen).^'^  These 
effects  suggest  strongly  the  presence  of  poisonous  substances  in  the 
blood  of  persons  or  animals  lacking  sufficient  thyroid  tissue. 

(3)  All  the  effects  of  thyroidectomy  are  more  marked  in  carnivor- 
ous animals  than  in  herbivora ;  indeed,  the  latter  may  be  able  to  live 

3  Concerning  tlie  influence  of  thyroid  on  skeletal  growth  see  Holmgren.  Xordiskt 
Med.  Arkiv,  1910  (43),  No.  2.  Literature  given  bv  Basingcr,  Arch.  Int.  31ed., 
1916   (17),  260. 

4  See  the  interesting  experiments  of  Oudernatsch  (Arch.  Entwickl.,  1J)12  (35), 
457;  Amer.  Jour.  Anat.,  1014  (15),  431:  Anat.  Record,  1017  (11),  357),  Avho 
found  that  feeding  thyroid  to  tadpoles  hastens  their  differentiation  but  checks 
growth. 

5  Jour,  of  Physiol.,  1005    (32),  150. 

6  Amer.  Jour.  Phvsiol.,  1004  (11),  351;  corroborated  by  Morse,  Jour.  Biol. 
Chem.,  1915   (22),  125. 

7  Jour.   Exper.   IVled.,    1000    (11),    665. 

sUnderliill  and  Saiki,  Jour.  Biol.  Chem.,   190S    (5),  225. 
8a  Hunter,  Quart.  Jour.  Phvsiol.,  1014    (8),  23. 
9Virchow's  Arch.,  1004    (176),  260. 
loVirchow's  Arch.,  1902    (170).  220. 


592  CHEMICAL    /'ATHOLOGY    OF    THE    DUCTLESS    GLAyDS 

in  fair  condition  for  several  years  without  a  thj-roid,^^  Administra- 
tion of  meat  to  thyroidcetomized  lierbivora  or  omnivora  causes  a  great 
increase  in  the  syinptoms,  while  tliyroideetomized  carnivora  do  much 
better  if  kept  without  meat.  Tims,  Hluui  ^-  found  that  thj-roidecto- 
mized  dogs,  which  were  doing  well  on  a  milk  diet,  developed  symptoms 
of  athyreosis  immediately  they  were  given  meat.  This  fact  has  been 
interpreted  as  indicating  that  toxic  materials  are  formed  from  meat 
in  tiie  intestinal  tract,  which  under  -normal  conditions  are  neutralized 
by  the  thyroid.  On  the  other  hand,  one  may  well  imagine  that  the  so- 
called  autointoxication  in  athyreosis  is  not  from  intestinal  putrefac- 
tion, but  is  due  to  the  products  of  incomplete  metabolism  of  proteins 
within  the  tissues,  which  are  destroyed  when  protein  metabolism  is 
normal,  but  not  when  the  metabolism-favoring  influence  of  the  thyroid 
is  wanting.  It  should  also  be  added  that  the  presence  of  specific 
poisonous  substances  in  the  blood  or  urine  of  thyroidcetomized  animals 
has  not  been  conclusively  established.^"' 

(4)  Reid  Hunt  ^*  found  that  mice  fed  thyroid  preparations  have  a 
greatly  increased  resistance  to  poisoning  by  aceto-nitrile ;  however,  this 
is  not  necessarily  nor  even  probably  a  direct  detoxication,  but  more 
likely  it  results  from  alterations  in  metabolism.  Rats  and  guinea 
pigs  behave  just  the  opposite,  showing  a  decreased  resistance  to  aceto- 
nitrile  after  being  fed  thyroid,  and  according  to  some  authors  morphine 
is  more  toxic  for  such  animals. ^"^^ 

"Whether  the  thyroid  exercises  its  detoxicating  effect,  assuming  that 
it  has  one,  by  a  direct  neutralizing  action  of  its  secretion  upon  the 
toxic  substances  in  the  blood  or  in  diverse  tissues,  or  indirectly  by 
stimulation  of  the  function  of  other  tissues  which  perform  the  de- 
toxication. or  in  part  locally  within  the  gland  itself,  is  an  unsettled 
problem.  In  relation  to  the  last-named  hypothesis  is  the  extreme  vas- 
cularity of  the  thyroid,  which,  according  to  Burton-Opitz  ^"  has  passed 
through  it  much  more  blood  in  proportion  to  its  weight  than  any 
other  gland.  Against  the  idea  of  a  local  detoxication  is  the  fact  that 
after  extir]iation  of  the  thyroid  all  abnormal  conditions  may  be  pre- 
vented l)y  i)r()pcr  administration  of  thyroid  substance. 

Biedl  summarizes  his  views  as  to  the  function  of  the  thyi'oid,   in 

11  I'arl  of  tlicse  results  may  be  due  to  tlie  faet  tliat  in  some  lierbivora  the 
parathyroids  are  so  far  separated  from  the  thyroid  that  they  are  not  ordinarily 
removed  in  thyroidectomy,  whereas  in  many  carnivora  eom]ilete  removal  of 
paratiiyroids  witli  tlie  thyroids  is  more  likely  to  be  aeeom])lished. 

12  Virchow's  Arch..   IHOO    (1(12),  .'575. 

i-i  Heinedi  ( Lo  Sperinientale,  1!)()2;  abst.  in  Cent.  f.  Path.,  litO.S  (14),  fi!)5) 
claims  tliat  tetanus  toxin  and  other  l)acterial  |)oisons,  when  injected  into  the 
thyroid  fjland,  are  harmless,  which  he  attrii)utes  to  a  neutralization  by  the 
colloid.  This  observation  is  discredited  bv  the  work  of  Basinger,  Jour.  Infect. 
Dis.,   1017    (20),   1.31. 

14  Jour.  Amer.  ^led.  Assoc,  1!H)7  (4!M,  240;  llyuicnic  Lul).  IbiU.,  1!>{)!I,  No.  47; 
Jour.  Pharmacol.,   1910    (2),   15. 

ir-See  Olds,   Amer.   Jour.    Physiol.,   1010    (2(1),  .Sr)4. 

I'UJuarl.  Jour.    Phvsiol..   101(>   (.'{).  207. 


cii i:\iisTin  OF  Tin:  riiynoiit  503 

tilt"  t'ollowiiiLj  staltMiiciit  :  "Tlic  tliyi'oid  is  a  secretory  orjraii  which 
(lischaf<i('s  its  secretion  eventually  into  the  blood,  in  tlie  form  of  an 
i()(lin-c()ntainin«i-  ])rotein.  Tliis  secretion  acts  as  a  horniont',  in  that 
it  niodilics  the  activities  of  remote  tissues.  As  far  as  we  now  know 
the  thyi-oid  secretion  i)lays  the  role  of  a  'disassiniilatory'  hormone,  in 
that  it  eauses  an  increased  disassimilation  and  increase  of  normal  activ- 
ity in  many  tissues.  This  effect  is  exemplified  by  the  au<,nncnted 
metabolism,  the  activity  of  the  heart  and  many  parts  of  the  symi)a- 
thetic  nei'vous  system,  and  of  a  series  of  internal  secretory  organs 
(adrenals,  liypophysis).  In  other  tissues  are  found  evidence  of  the 
action  of  an  inhibiting'  and  assimilatory  hormone,  as  shown  in  the  in- 
crease in  growth  of  bone,  development  of  the  sex  glands,  and  de- 
creased internal  secretion  of  the  pancreas." 

CHEMISTRY  OF  THE  THYROID  i" 

Whether  the  function  of  the  tliyroid  is  the  neutralization  of  toxic 
substances,  or  a  complementary  action  upon  intracellular  metabolism, 
there  can  be  little  question  that  it  owes  its  action  to  constituents  of 
its  specific  secretion,  the  colloid.^'''  Furthermore,  the  chief,  if  not  the 
sole,  active  ingredient  of  the  colloid  is  the  iodin-containing  substance 
first  discovered  by  Baumann  in  1896,  and  called  by  him  thyroiodin 
(or  iodothyrein) ?''^ 

The  chemical  nature  of  thyroid  colloid  has  been  studied  particularly 
b}-  Oswald.'*  lie  found  that  all  the  iodin  of  the  thyroid  is  dissolved 
out  in  physiological  salt  solution,  and  that  none  of  it  is  present  in  an 
inorganic  form.  In  the  salt  solution  extract  are  two  protein  bodies ; 
one,  precipitated  by  half  saturation  with  ammonium  sulphate,  con- 
tains all  the  iodin,  and  seems  to  be  a  globulin ;  it  resembles  myosin 
in  being  precipitated  by  weak  acids,  and  it  contains  an  easily  separated 
carbohydrate  group.  The  other,  precipitated  by  saturation  with  am- 
monium sulphate  (exact  limits  of  precipitation  are  between  6.4  and 
8.2  tenths  saturation),  is  a  nucleoprotein,  containing  0.16  per  cent, 
phosphorus,  but  no  iodin. 

The  iodin-containing  protein,  called  fhyreoglohulin,  constitutes  one- 
fourth  to  one-half  the  dry  weight  of  the  gland,  and  seems  to  be  the 
sole  active  constituent  of  the  colloid ;  at  least,  its  administration  to 
animals  has  the  same  physiological  effects  as  does  the  entire  colloid 
(great  increase  in  the  urea  elimination  and  decrease  in  blood  pressure 

17  C4ood  reviews  are  triven  bv  Rahel  Hirsch.  Handb.  d.  Biocbem.,  1900,  III  (,), 
271;  and  A.  Kocber,  Vircbow's  Arch..  1912    (208),  86. 

i"a  Beyond  tlie  cliaracteristio  roHoid  se  -retion  product,  tlie  tbyroid  presents  no 
cbemical  features  of  interest;  it  ditTers  from  tbe  otlier  endocrine  glands  in  l)ein<i 
poor  in  lipoids  (  Fenger,  Jour.  Biol.  Clieni.,  IflKi   (27),  .SO."?). 

17b  Iodin  is  present  in  tbe  tbvroid  of  all  species,  most  in  marine  fornis  (Cam- 
eron, Jour.  Biol.  Cbem..  1014   (Ki).  4(i.');   Biocbem.  .Tour.,  1014   (7).  406). 

1^  His  work  is  reviewed  in  liis  dissertation,  ''Die  cbem.  BescbafTenheit  nnd  die 
Function    dcr    Scbilddruse,"    Strassburg,    1000;    also    see    Vircbow's    Arcb.,    1002 
(169),  444. 
38 


59-4  CHEMICAL    PATHOLOGY    OF    THE    DUCTLESS    GLAXDS 

ill  animals,  curative  effect  on  myxedematous  patients),  whereas  the 
nucleoprotein  is  witliout  these  effects.  Analysis  of  the  thyreoglobulin 
from  various  animals  has  shown  it  to  be  of  quite  constant  quantitative 
composition  except  for  the  iodin,  which  may  vary  greatly  in  amount. 
Normal  human  thyreoglobulin  (from  persons  living  in  non-goitrous 
districts)  had  the  following  percentage  composition: 

C  =  51. So,    H  =  0.88,    X  =  15.49,    I  =  0.34,    S  =  l.SG. 

Th,yreoglobulin  from  goitrous  districts  contains  much  less  iodin 
(0.18-0.19  per  cent.l,  and  from  calves  born  with  goiters  a  thyreo- 
globulin was  obtained  that  agreed  in  all  respects  with  nonnal  thyreo- 
globulin, except  that  it  contained  no  iodin  at  all.  On  the  other  hand, 
administration  of  iodides  to  patients  causes  the  thyreoglobulin  to 
become  rich  in  organically  bound  iodin.^**  From  these  facts  Oswald 
believes  that  the  thyreoglobulin,  as  first  secreted  by  the  glandular  epi- 
thelium, is  free  from  iodin,  and  that  it  combines  later  with  iodin  from 
the  circulating  blood.  As  yet  it  has  not  been  ascertained  how  the 
iodin  is  bound  to  the  protein.  It  is  well  known  that  large  amounts 
of  iodin  can  be  introduced  into  the  protein  molecule,  apparently 
through  its  substitution  for  hydrogen  in  the  aromatic  radicals  (tyro- 
sine, phenylalanine,  etc.)  Thyreoglobulin  is  not,  however,  simply  an 
iodized  protein,  for  the  iodized  proteins  that  can  be  artificially  pre- 
pared do  not  possess  the  physiological  activity  of  the  thyreoglobulin ; 
furthermore,  the  saturated  iodized  proteins  contain  generally  from  5 
to  12  per  cent,  of  iodin,  as  contrasted  with  the  0.8  to  0.8  per  cent,  of 
thyreoglobulin.  Oswald  has  shown  that  in  thyreoglobulin  the  iodin 
is  not  bound  to  tyrosine,  since  this  can  be  removed  by  tryptic  digestion 
without  decreasing  the  amount  of  iodin  in  the  rest  of  the  molecule ; 
possibly  the  iodin  is  bound  to  phenylalanine.-"  F.  C.  Koch  -^  finds  that 
the  full  activity  of  the  gland  is  contained  in  the  thyreoglobulin,  and 
also  in  the  metaprotein  fraction  of  this  globulin,  while  simpler  cleav- 
age products  show  less  and  less  activity  per  unit  of  iodin  content. 
Tie  could  find  no  thyroid  activity  in  iodin  compounds  of  histidine,  and 
di-iodotyrosine  was  found  inactive  by  Strouse  and  Voegtlin.-- 

The  remarkable  influence  of  the  thyroid  on  the  development  of  tad- 
poles (Gudernatsch  *)  is  exhibited  by  the  thyreoglobulin,  but  not  by 
any  other  iodin  compound  that  has  been  tried,  except  possibly  by 
iodized  blood  protein  (Morse  ^^'').  Abderhalden,^-"'  however,  found 
lliat  if  th^-roid  was  digested  until  nearly  or  (piite  biuret-free  and  then 

10  Nape!  and  Tfoos  (Arch.  f.  Anat.  u.  Physiol.,  1002,  p.  2(17)  foiiiul  t'lat  ad- 
ministration of  bromidos  had  no  effoct  upon  the  amount  of  iodin  in  tlu>  thyroid, 
and  no  storaf^o  of  bromin  ta'^o*  T-'aoe.  Administration  of  pilocarpine  docs  not. 
increase  ilie  amount  of  iodin  iTi  tlic  thyroid. 

20  Niirn1)cri:.  llofincisicr's  Bcitr..  1007    (10),  125. 

•■;i  Jour.   l?iol.   Cliem.,    101:5    (14),    101. 

-'-•Jour.  Pliarni.  and  F\p.  Ther.,  1010    (1).  123. 

1!'^' Jour.  l?iol.  C'hcm.,  1015   (10),  421. 

i'"'Arcli.  ^'cs.  Phvs..   1015    (102),  00. 


CHEMISTRY  OF  Till-:  THYUOin  595 

dialyzcd,  tlie  active  suhstaiiee  diffused  thnnimh  the  membrane,  indi- 
cating that  it  is  not  a  colh)idal  complex,  and  that  if  derived  from  the 
proteins  it  must  be  an  iodized  amino-acid  or  some  related  compound. 
This  observation  is  to  be  expected  in  view  of  the  fact  that  thyroid  pro- 
duces its  effects  when  fed  and  is  presumably  hydrolyzed  before  ab- 
sorption. Lenhart  ^'-"^  considers  the  effect  of  thyroid  on  tadpoles  to  be 
merely  an  expression  of  the  (>oneral  stinndatio]i  of  metabolism,  rather 
than  a  specitic  effect  on  differentiation.""^ 

liy  decomposing  thyreoglobulin  by  boiling  with  30  per  cent,  sul- 
phuric acid,  a  body  is  obtained  containing  as  high  as  14.5  per  cent,  of 
iodin ;  this  is  the  tJnjroiodin  of  Baumann,  which  gives  no  biuret  re- 
action, yet  is  physiologically  active.  The  stability  of  this  active  con- 
stituent of  the  thyreoglobulin  explains  the  successful  administration 
of  thyroid  preparations  by  mouth.  It  appears  to  be  absorbed  un- 
changed and,  unless  enormous  doses  are  given,  none  appears  in  the 
urine  (Oswald -■'').  Long-continued  digestion  with  trypsin,  or  auto- 
lysis of  thyroid  glands,  causes  a  complete  splitting-out  of  the  iodin. 
One  part  of  the  iodin  seems  to  be  more  firmly  bound  than  the  rest. 

Kendall  ~*  has  isolated  from  the  thyroid,  after  alkaline  hydrolysis,  a 
crystalline  compound  containing  60  per  cent,  of  iodin,  which  he  sug- 
gests may  be  di-iodo-di-hydroxy-indole.  This  is  highly  active,  causing 
rapid  pulse,  nervous  irritability,  and  increased  metabolism.  It  does 
not  contain  all  the  iodin  of  the  thyroid,  but  the  nature  of  the  other 
iodin  compounds  is  unknown  beyond  the  observation  that  they  have 
no  appreciable  effects  on  normal  persons  but  greatly  improve  the  con- 
dition of  cretins.  A  small  amount  of  the  iodin  may  exist  as  inorganic 
and  lipoid  compounds.-*^  When  fed  to  tadpoles,  Kendall's  active 
principle  produces  the  characteristic  thyroid  effect."*'' 

The  amount  of  iodin  in  the  thyroid  is  greatest  in  middle  age,  greater 
in  females  than  in  males,  and  it  is  decreased  in  acute  infectious  dis- 
eases and  in  tuberculosis,  alcoholism,  and  circulatory  disturbances 
(Aeschbacher).'^ 

The  thyroid  is  very  rich  in  lipase,  catalase  and  peroxidase;  ex- 
tirpation is  followed  by  a  decrease  in  these  enzymes  in  the  blood, 
while  thyroid  feeding  increases  them  as  well  as  the  antitrypsin 
( Juschtschenko)  .^® 

The  physiological  activity  of  thyroid  preparations,  according  to 
nearly  all  investigators,  is  in  direct  proportion  to  the  iodin  content,-'' 

19c  Jour.  Exp.  Med.,  1015    (22),  739. 

iMSee  also  Kahn,  Arch.  (jes.  Physiol.,  1016   (163),  384. 

23  Arch.  exp.  Path.  u.  Pharm.,  1910   (63).  263. 

24  Jour.  Amer.  Med.  Assoc,  1015   (64),  2042;  Jour.  Biol.  Cheni.,  1915   (20),  501. 
2-ta  Blum  and  Griitzner,  Zeit.  physio].  Cheni.,  1914    (91),  400. 

24b  Rogoff  and  ^Marine,  Jour.  Pharmacol.,  1910    (9),  57. 

25  :Mit't.  a.  d.  Grenzgeb.  med.  u.  C'hir.,  1905  (15),  209;  Pellegrini,  Arch.  sci. 
med.,  1915   (39),  276. 

26Biochem.  Zeit.,  1910  (25),  49;  Zeit.  phvsiol.  Chem.,  1911   (75),  141. 

27  Fonio,  Mitt.  Grenz.  Med.  u.   Chir.,   1911    (24),  123;   Frey,  ibid.,   1914    (28), 


596  CHEMICAL    PATHOLOGY    OF    THE    DUCTLESS    GLAyOS 

which  is  the  best  of  evidence  that  the  formation  of  this  compound  is 
one  of  the  chief  functions  of  the  gland,  and  that  the  iodin  in  the  thy- 
roid is  not  merely  stored  there  as  an  undesirable  foreign  substance 
like  copper  in  the  liver.  The  selective  deposition  of  iodin  in  the  thy- 
roid is  remarkable,  and  when  iodin  is  fed  to  animals  it  is  stored  very 
rapidlj'  in  the  thyroid,  bvit  it  seems  to  require  several  hours  before  the 
active  growth-modifying  hormone  is  formed.-^"  INIarine  and  Lenhart  -^ 
find  that  the  normal  human  gland  contains  an  average  of  0.4  mg.  of 
iodin  per  gram  of  fresh  weight  (2.17  mg.  per  gram  of  dry  weight),  be- 
ing less  than  that  of  domestic  animals  in  the  same  part  of  the  country. 
These  figures  agree  closely  with  tliose  obtained  in  thyroids  from  various 
parts  of  America  by  Wells--'  (2.10  mg.  per  gram  dry  weight).  They 
found,  as  Oswald  and  Kocher  also  have,  that  the  amount  of  iodin  varies 
directly  with  the  amount  of  colloid,  being  decreased  when  cellular  hy- 
perplasia is  present,  in  direct  proportion  to  the  amount  of  hyper- 
plasia, and  administration  of  iodin  causes  a  reduction  in  the  In'per- 
plasia  and  a  return  to  the  colloid  type  of  gland,  while  the  iodin  is  de- 
posited in  the  gland.  Kocher,  however,  disputes  the  regularity  of  the 
variation  of  iodin  and  colloid  content,  stating  that  it  is  especially  the 
concentrated  follicle  contents  which  hold  the  iodin.  Seidell  and  Fen- 
ger  ^°  have  found  a  marked  seasonal  variation  in  the  thyroid  iodin  of 
animals,  there  being  about  three  times  as  much  between  June  and  No- 
vember as  between  December  and  May.""''  In  man  it  has  been  found 
that  before  birth  the  thyroid  of  the  fetus  contains  little  or  no  iodin, 
but  in  domestic  animals  the  fetal  glands  contain  not  a  little  iodin 
(Fenger).^^  The  cells  of  the  gland  contain  very  little  iodin  (A. 
Kocher).  Extracts  of  the  thyroid  have  little  effect  on  the  blood  pres- 
sure, except  for  an  alcohol-soluble  fraction,  poor  in  iodin,  which  is  a 
depressor.^^^ 

Wasting  diseases  are  associated  with  a  considerable  decrease  in  the 
size  of  the  thyroid  and  the  amount  of  colloid,  and  with  this  a  decrease 
in  the  iodin ;  especially  is  this  true  of  tuberculosis.^-  Patients  or  ani- 
mals to  whom  iodin  compounds  are  administered  deposit  it  in  the 
thyroid  in  large  amounts,  especially  if  the  gland  is  previously  de- 
fective in  iodin,  and  at  times  there  results  even  an  acute  thyroiditis 
from  the  iodin  administration.^^     Iodides  are   said   to   increase   the 

340;  Hunt.  Jour.  Amer.  Med.  Assoc,  1907  (49),  1323;  and  Jour.  Pharm.  and  oxp. 
Therap.,  1910   (2),  15. 

27a  Marine  and  l^ogofi",  Jour.  Pliarm.,  191G    (9),  1. 

28  Arch.   Int.   Med.,    1909    (4),  440. 

29  Jour.  Amer.  :Med.  Assw.,   1897    (29),  897. 

30  .Tour.  P.iol.  Clieni.,  1913    (13),  r,]l. 

30a  \'alual)k'  fifjures  on  tlic  iodin  content  of  foods  are  given  bv  Forhes  ct  at., 
Bullet.  Oliio  Agric.  Exi)t.  Stotion,  June,  1910.  No.  299. 

31  Jour.  Piol.  Cliem.,  1912   (11),  489;    1912   (12),  .5;);   1913    (14),  397. 
3ia  Fawcett  et  al.,  Amer.  .Four.  Pliysiol.,  1915  (.3(5),  113. 

32  See  Vitrev  and  Hiraud,  Conijjt.'Rend.  Soc.  Biol.,  1908   (65),  405. 

33  See  Mendel,  .Med.   Klinik,  19()(;    (2),  833. 


77//;    l'Mr\TII  )!,'() IDS 


597 


ainoiiiit  of  tliyi-('()<i-l()l)uliii  itself  (Wiener).'"  The  variation  in  iodin 
content  under  various  conditions  is  given  in  tlie  following  table  from 
Jolin,^''  his  tigures  for  normal  glands  being  somewhat  lower  than 
found  ill  America. 


Number  and  Condition  of  Glands 

Dry  Wt. 

Gms 

Mg.  Iodin 
Per  Gm 

Total 
Iodin 

152   glands   from   piMsuiis  over   10  yrs.  oUl    (44  not 

normal )                

7  04 

1  03 

11.20 
0.145 

28  glands  from  eliildren   (1  mon.  to  10  yrs. ) 

0.54 

0.28 

108  normal  glands  from  adults  only   (both  sexes) 

5.38 

1.56 

8.05 

(J7  normal  glands  from  adults   (males) 

5.07 

1.50 

7.83 

41   norn:al  glands  from  adults    (females) 

5.no 

1.55 

8.40 

38  glands  from  chronic  di.-eases 

4.29 

1.90 

7.81 

2!)  glands  fiom  acute  diseases 

5.54 

1.47 

8.11 

21    glands   from   suddi'n  death 

6.88 

1.29 

8.45 

lit  glani's   slinwing  marked  goiter 

23.09 

1.09 

26.49 

25  colloid-rich  glands 

8.25 

2.24 

18.20 

34  glands  from  persons  receiving  iodin 

5.79 

2.56 

15.06 

THE  PARATHYROIDS  36 

"Whether  the  thyroid  has  anj-  other  function  than  the  formation 
of  thyroiodin  is  as  yet  unknown.  ^INIany  claim  that  the  th3'reoglobulin 
does  not  produce  the  same  physiologic  and  therapeutic  effects  as  does 
the  entire  gland  substance,  but  even  that  is  not  definitely  decided. 
Furthermore,  much  of  the  difficulty  comes  from  the  failure  of  earlier 
observers  to  distinguish  between  the  effects  produced  by  the  para- 
thyroid glands  and  those  due  to  the  thyroid  itself.  The  parathyroids 
were  originally  considered  as  but  a  form  of  undeveloped  accessory- 
thyroids  (a  view  still  held  by  some),  but  they  are  now  generally  be- 
lieved to  be  independent  organs  of  fully  as  great  importance  as  the 
thyroid.  Their  independence  is  conclusively  shown  by  the  cases  of 
cretinoid  children  in  Avhom  the  thyroid  proper  has  failed  to  develop, 
while  the  parathyroids  are  found  to  be  normal,^'  thus  proving  their 
distinct  origin,  the  inability  of  parathyroid  tissue  to  change  into  thy- 
roid ti.ssue,^*  and  their  inability  to  prevent  the  changes  of  cretinism.^* 
Parathyroids  contain  no  appreciable  amounts  of  iodin  (Estes  and 
Cecil V^"  although  14  per  cent,  of  parathyroids  obtained  at  autopsy 
contain  a  colloid  material  (Thompson  and  Harris).*^  Glycogen  is 
demonstrable  in  the  epithelium.     To  their  removal  are  ascribed  by 

34  Arch.  exp.  Path.  u.  Pliarm.,  1909   (01).  297. 

33  Festsehr.  f.  O.  Ilammarsten,  Upsala  Lakarefiiren.  Fr>rh.,  190G.  XI,  Sujipl. 

s"  An  excellent  review  of  tliis  subject  is  given  by  Thom})son  in  "The  Surgery 
and  PathologT  of  the  Thyroid  and  Parathyroid  Giands."  bv  A.  J.  Ochsner  and 
R.  L.  Thompson,  St.  Louis,  1910.  See  also  MacCallum,  Ergeb.  inn.  :\Ied.,  1913 
(11),  509. 

37  Roussy  and  C'lunet,  Compt.  Rend.  Soc.  Biol. 

38  See  Edmunds,  Jour.  Path,  and  Pact.,  1910 

39  See  ^MacCallum,  Johns  Hopkins  Hosp.  Pull., 

io  fhid.,   1907    (18),  331:    also  Cameron,  J.mr.   Biol.  Clu-in..    1914    (10),   465. 
41  Amer.  Jour.  Med.  Sci.,  1908   (19),  135. 


1910   (68),  818. 
14).  288. 
1907    (IS).  341. 
Biol.  Cliein..    1914 


598  CHEMICAL    PATHOLOGY    OF    THE    DUCTLESS    GLAXDS 

many  investigators  the  acute  manifestations  of  athyreosis,  while  the 
more  chronic  changes  of  myxedema  are  attributed  to  the  loss  of  the 
thyroid.  MacCallum's  studies  support  this  \dew,  for  he  found  the 
results  of  parathyroidectomy  in  dogs  very  different  from  the  results 
of  thyroidectomy.  The  most  prominent  symptoms  are  muscular 
twitchings,  gradually  passing  into  tetanic  spasms,  and  due  to  nervous 
impulse  rather  than  to  muscular  changes,  since  they  did  not  appear 
in  muscles  from  which  the  nerve-supply  has  been  cut  off.  Trismus, 
protrusion  of  the  eyes,  and  rapid  respiration  without  cyanosis  {i.  e.. 
air  hunger)  were  also  observed,  and  death  usually  resulted  from  ex- 
haustion. Apparently  these  sj^mptoms  are  due  to  some  toxic  sub- 
stance which,  accumulates  on  account  of  the  absence  of  the  parathy- 
roids, for  it  was  found  that  simply  diluting  the  dog's  blood  by  with- 
drawing part  of  it,  and  injecting  a  corresponding  amount  of  salt  so- 
lution, caused  a  temporary  cessation  of  the  tetanic  symptoms ;  and 
injections  of  emulsions  of  parathyroid  checked  the  sj'mptoms  for 
some  time,  presumably  through  neutralizing  the  hypothetical  poisons. 
Degenerative  changes  that  were  observed  in  the  cerebral  ganglion- 
cells  also  favor  the  view  that  some  unneutralized  toxin  is  responsible 
for  the  symptoms  following  parathyroidectomy,  and  W.  F.  Koch  *^^ 
found  that  the  urine  of  parathyroidectomized  animals  contains  an 
abundance  of  toxic  bases,  especially  methyl  cyanamide.  On  the  other 
hand,  profound  mental  symptoms  and  insomnia  have  resulted  from 
feeding  parathyroid  to  man.*^'' 

The  metabolism  after  parathyroid ectomii  may  show  tlie  following  ohaiities :  -12 
There  is  a  reduction  in  the  assimilation  limit  for  carbohydrates  (Hirsch,  Under- 
hill  42a  and  others).  Concernino;  inorganic  metabolism  there  is  disasjreement,  for 
while  ilacCalliim  and  Voegtlin  *^  found  an  increased  elimination  of  calciinn  and 
a  loss  of  tlie  same  element  from  the  blood  and  brain  (which  they  would  make 
responsible  for  the  increased  nervous  irritability),  Cooke  found  no  such  loss  of 
calciiun,^3a  but  she  did  find  an  increased  urinary  excretion  of  magnesiiun.  Ac- 
cording to  most  observers,  nitrogenous  metabolism  is  altered  as  shown  by  the  in- 
creased excretion  of  nitrogen,  and  especially  of  anunonia,  which  suggests  tlie  ex- 
istence of  an  acidosis.  Greenwald  **  found  increased  anunonia  less  conspicuous 
than  increased  undetermined  nitrogen  and  sul])hur,  and  decreased  ])hos])horus 
excretion.  There  may  occur  an  increase  in  the  bases  of  the  blo;id  (alkalosis) 
which  disappears  under  the  acidosis  that  results  from  tetany. ■♦^i> 

In  view  of  the  conflicting  facts,  the  theory  that  the  increased  irritability  and 
spasm  of  tetany  result  from  hyi)ocalcification  of  tlie  nerve  tissue  is  at  present 
unproved.  Calcium  does  diminish  nervous  irrital)ility,  as  shown  by  J.  Loeb, 
and  hence  when  administered  it  may  favorably  intlueiice  the  symptoms  of  tetany 
para.tli\re()])riva.,  liut  Ihis  does  not  establish  tlie  tlieory.  Tliat  luunerous  experi- 
menters have  been  able  to  stop  tliese  symptoms,  both  in  man  antl  animals,  by 
feeding  of  parathyroid,*5  or  parathjroid  nucleoprotein,  establishes  the  relation- 

4ia.rour.  Uiol.  Chem.,  11)1.3   (1,5).  4.3;  Jour.  Lab.  Clin.  :\led.,  lOK]   (1),  2!1!). 

41b  Morris,  .lour.  F.ab.  Clin.  Med.,  IDl.'i  ( 1 ) ,  2(i. 

■J 2  Sec  review  Ijv  Cooke,  .\mer.  dour.  :Med.  Sci..   1010    (UOi,  404. 

42a  .Tour.  Biol.  Cliem.,  1014    (IS),  87. 

43  Jour.  K\p.  Med.,  1009   (11),  118;  101.3   (IS).  CIS. 

43a  See  also  IJergeim,  Stewart  and  Hawk,  Jour.  Kxp.  :Med.,  1014   (20),  225. 

44Amer.  Jour.  I'liysiol.,   1011    (2S),  10.3;  Jour.  Biol.  Chem.,  1013    (14),  3()3. 

44a  Wilson,  Stearns  and  'i'hurlow,  .lour.  Biol.  Chem..   I!)!')    (23).  SO,    123. 

45  See  Ilalsted,  Amer.  .lour.  ISied    Sei.,   1007    n34),   1. 


cnHuisTJi'Y  OF  (!<UTi:n  599 

ship  of  this  <rhui(l  to  the  totatiy,  hut  not  tlio  cah'iuiii  deprivation  liypothosis.  A 
critique  of  tiiis  liyi)otiu'sis  hy  lU'ikch'y  and  IJeebo  ■""'  hrings  out  the  foih^winf^ 
points:  Strontium,  inaj^ncsiuni  and  harinni  liavo  tiie  same  elVect  in  tetany  as 
fak'iiim,  wiiereas  severe  eah-iuni  loss  in  diahetie  acidosis  does  not  cause  tetany. 
Tlie  fact  tiiat  Ideedinj,'  reduces  tiie  symptoms  is  ai^^ainst  tiie  cahdum  de])rivation 
theory  and  supports  the  intoxication  tiieor.y.  ^^'icne^  47  even  states  tiiat  it  is 
possible  to  secure  an  antitoxin  for  tiie  poison  of  tetany  thyreopriva  hy  immuniz- 
ing with  the  serum  of  animals  in  tetany.  On  the  otlier  hand,  the  marked  clianj;es 
in  dentition  and  l)one  repair  observed  in  paratliyroidectomized  animals  liy  Krd- 
heim  -^s  indicate  an  abnormality  in  calcium  metabolism,  whicli,  however,  mif;ht  be 
secondary  to  an  intoxication.  Also,  in  osteomalacia  and  osteoporosis  the  para- 
thyroids are  said  to  siiow  hyperjilasia,^''  and  1  lowland  and  Marriott  have  found  a 
definite  decrease  in  the  calcium  of  the  blood  in  human  tetany  and  in  paratliyroid- 
ectomized doi:s.i'''i 

MacCallum  •'^^"  has  fcmiul  evidence  that  in  paratliyroidectomized  dogs  the  blood 
contains  something  which  greatly  increases  the  irritability  of  tiie  nerves,  possibly 
by  abstracting  calcium  from  the  tissues.  Removal  of  calcium  from  the  blood  by 
dialysis  results  in  nerve  hyperexcitability  resembling  that  seen  in  tetany. 

Cooke  states  that  the  metabolic  changes  precede,  and  presumably  incite  tlie 
tetany.  Im])lantation  of  parathyroid  tissue  in  persons  with  tetany  parathyreo- 
priva  has  been  successful  in  removing  symptoms  in  a  few  cases. 5i  The  relation 
of  the  parathyroids  to  tetany  of  infants  is  not  so  well  established."'^  altliough 
several  observers  have  found  hemorrhages  in  the  paratliyroids  in  these  cases. 
Some  cases  of  "gastric  tetany''  have  improved  under  parathyroid  feeding,  which 
is  also  said  to  be  beneficial  in  paralysis  agitans,^-^  although  there  seems  to  be 
no  anatomic  basis  for  assuming  a  parathyroid  deficiency  in  this  disease. 

CHEMISTRY  OF  GOITER 

In  connection  with  his  earliest  studies  of  thyroiodin,  Baumann  ob- 
served a  great  difference  in  the  amount  of  iodin  in  the  thyroid  glands 
of  normal  individttals  living  in  goitrous  districts,  as  compared  with 
those  living  in  non-goitrous  districts.  Thus  in  Freiburg,  a  goitrous 
district,  the  average  weight  of  the  dried  thyroid  was  8.2  grams,  each 
gram  containing  0.33  mg.  of  iodin,  a  total  of  2.5  mg.  of  iodin  to  each 
gland.  Glands  from  Hamburg  averaged  4.6  gm.  in  weight,  contain- 
ing 0.83  mg.  of  iodin  per  gram,  a  total  of  3.83  mg.  per  gland.  Berlin 
glands  weighed  7.4  grams,  and  contained  0.9  mg.  of  iodin  per  gram, 
or  a  total  of  6.6  mg.  of  iodin  per  gland.  Both  of  the  last-named  cities 
are  in  districts  where  goiter  is  not  endemic.  The  thyroids  of  young 
children  show  the  same  relative  paucity  of  iodin  in  goitrous  districts, 
as  compared  with  non-goitrous  districts.  Wells  ^^  found  that  the  thy- 
roids throughout  the  United  States  contain  even  larger  amounts  of 
iodin  than  the  Berlin  glands,  averaging  10  to  12  mg.  per  gland,  agree- 
ing with  the  fact  that  goiter  is  comparatively  rare  in  this  country.'^'* 

*fiJour.  Med.  Res.,   1000    (20),  140. 

47  Pfliiger's  Arch.,   inin    fl.3G).  107. 

4s  Frankfurter  Zeit.  Rathol..  1011    (7),  175. 

40Todyo,  Frankf.  Zeit.  Pathol.,   1012    (10),  210. 

4na  Trans.  Amer.  Fed.  Soc,  Vol.  28.  p.  202. 

noVerh.  Deut.  Path.  Cies..  1012    (15).  206:   Jour.  Exp.  Med.,  1914    (20),   140. 

51  Danielsen,  Beit.  klin.  Chir.,  1010   (06),  85. 

52  See  Haberfeld,  Virchow's  Arch.,  1011    (203),  282. 
52a  Berkeley,  Med.  Record,  1016    (00),  105. 

53  Jour.  Amer.  Med.  Assoc,   1807    (20),  807. 

54  It  is  probable,  in  view  of  the  higher  results  obtained  by  later  analyses,  that 
the  results  of  Baumann  and  of  Monery  are  somewhat  too  low. 


600  CHEMICAL    PATHOLOGY    OF    TIN-:    DICTLE.^H    GLAXDS 

Mouery  '•'  has  found  for  France,  as  Baumann  did  for  Germany,  that 
the  amount  of  iodin  contained  in  the  glands  of  normal  individuals  is 
in  inverse  proportion  to  the  fre(iuency  of  g'oiter  in  districts  in  which 
they  live.  Oswald,  and  also  Aeschbacher,^*^  however,  state  that  normal 
thyroids  in  goitrous  districts  contain  more  iodin  than  thyroids  from 
goiter-free  districts. 

Chemical  analyses  of  goiters  have  given  extremely  variable  results, 
which  are  found  to  depend  upon  the  histological  type  of  the  goiter. 
Baumann  found  th^t  in  a  series  of  twelve  cases  of  goiter,  in  which 
the  average  dry  weight  was  32  grams,  the  amount  of  iodin  in  each 
gram  was  but  0.09  mg.,  but  the  total  amount,  2.6  mg..  was  about  the 
same  as  in  normal  glands  of  the  same  goitrous  district.  However, 
in  two  goiters  large  amounts  of  iodin  were  found,  namely,  17.5  mg. 
and  31.5  mg.  "Wells  found  that  the  amount  of  iodin  depended  upon 
the  structure,  for  two  hyperplastic  goiters  contained  respectively 
8.23  and  8.3  mg.  of  iodin,  or  about  the  amount  normal  for  thyroids 
in  this  country,  whereas  two  colloid  goiters  contained  53.16  and  24.59 
mg.  of  iodin.  This  is  corroborated  by  the  more  extensive  studies  of 
^Marine  and  his  co-workers,  who  have  found  the  proportion  of  iodin 
low  in  all  glands  showing  epithelial  hyperplasia,  but  high  in  colloid 
goiters.""  Administration  of  iodin  causes  a  reversion  of  the  hyper- 
plastic to  the  colloid  tj'pe  of  gland,  while  deprivation  of  iodin  causes 
hyperplasia.  Presumably,  therefore,  during  the  active  growth  of  a 
goiter  the  iodin  is  low,  but  in  the  quiescent  colloid  state  it  is  high. 
The  physiological  activity  of  colloid  or  other  preparations  from  goi- 
ters is  found  to  be  quite  the  same  as  that  from  normal  glands,  vary- 
ing in  direct  proportion  to  the  iodin  content."^^  In  an  adenomatous 
goiter,  in  the  new  growth,  AVells  found  1.98  mg.  of  iodin  per  gram, 
Mhile  the  rest  of  the  gland  contained  but  0.8  mg. ;  the  total  amount  of 
iodin  was  9.26  mg.,  or  the  same  quantity  as  found  in  normal  glands. 
In  nine  fetal  adenomas  ^larine  and  Lenhart  found  iodin  in  eight  in 
amounts  averaging  0.174  mg.  per  gram  of  dry  weight.  However,  when 
iodin  is  given  to  persons  with  thyroid  adenomas  the  tumor  tissue  does 
not  take  up  the  iodin  to  the  same  extent  that  the  normal  gland  tissue 
does. 

Oswald  found  that  colloid  goiters  contain  a  thyreoglobulin  that  is 
relatively  very  poor  in  iodin;  in  goitrous  calves  the  thyreoglobulin 
contained  no  iodin;  in  human  goiters  it  contained  but  0.07  to  0.19  per 
cent,  of  iodin,  as  against  a  normal  proportion  of  0.34  per  cent.  Ad- 
ministration of  iodides  to  a  goitrous  patient  caused  a  rise  in  the  pro- 
portion of  iodin  in  the  colloid  to  0.51  ])er  cent.,  showing  that  in  colloid 
goiters  in  goitrous  districts  the  thyi-eoglobuliii  is  probably  jioor  in 
iodin  because  of  a  lack  of  iodin  for  it  to  unite  with,  and  not  because 

•''-'.Tour.  Pliarm.  ot  Ciiiiii.,  1004    («).■)),  2SS. 

■"'I  Mitt.  a.  d.  f)ronz<roh.  Mod.  u.  Cliir..  ino.')   (IT)),  2{]9. 

"Arch.  Int.  :Med.,  1!)08    (1).  34!l:    lOOO    (.3).  fiti;    IflOO    (4).  440. 

58  See  Fonio,  Mitt.  Grenz.  :Med.  u.  <liir..  1011   (24),  123. 


.1/  yxKDi:  1/ 1    \\i)  ('inrri  \  is  1/    .  601 

it  is  of  ail  al)ii(»rinal  nature  that  ])i'events  its  eluMiiical  coinbiuatioii 
with  iodin.''"  Possibly  this  explains  the  greater  iodiii  content  ob- 
served in  colh)id  goiters  in  the  United  States  as  compared  with  eoUoid 
goiters  observed  in  goitrous  districts.  In  general,  Oswald ""  found 
the  amount  of  iodin  to  vary  with  the  amount  of  colloid  in  the  goiters, 
although  occasionally  goiters  with  exceptionally  large  amounts  of  iodin 
were  found,  and  the  proportion  of  iodin  is  not  usually  so  great  when 
the  amount  of  colloid  is  very  large.  Simple  hj'perplastic  goiters  he 
found  i)oor  in  iodin,  or  free  from  it  if  they  contained  no  colloid; 
however,  they  were  found  to  contain  a  thyreoglobulin  tyjiical  in  all 
respects  except  an  absence  of  iodin.  Presumably  in  such  goiters  the 
little  thyroiodin  present  is  contained  in  the  parenchymatous  cells. 
The  physiological  activity  of  thyreoglobulin  obtained  from  goiters  was 
found  to  be  the  same  as  that  from  normal  glands,  exce])t  that  it  was 
weaker  in  direct  proportion  to  the  amount  of  iodin  it  contained,  and, 
therefore,  when  iodin-free  it  w^as  without  effect.  In  colloid  goiters 
the  greater  part  of  the  weight  of  the  gland,  three-fourths  or  more,  is 
made  up  of  this  colloid-poor  thyreoglobulin.  The  fluid  contents  of 
cystic  goiters  may  be  free  from  iodin,  but  if  they  contain  much  colloid, 
iodin  will  be  found,  and  Rositzky  "^  found  0.193  mg.  of  iodin  in  20  c.c. 
of  the  jelly-like  contents  of  a  thyroid  cyst. 

It  has  been  frequently  suggested  that  the  cause  of  endemic  goiter 
is  a  deficiency  in  the  iodin  in  the  food,  or  in  the  drinking-water,  or  in 
the  air  of  the  goitrous  district.  This  is  supported  by  the  relative  in- 
frequency  of  endemic  goiter  in  districts  on  the  sea-coasts,  where  the 
iodin-containing  sea-water  is  sprayed  through  the  air,  and  where  the 
inhabitants  eat  largely  of  sea-foods.  However,  there  are  many  ex- 
ceptions, and  it  cannot  be  said  that  this  hypothesis  of  the  etiology  of 
goiter  rests  on  satisfactory  evidence,  particularly  in  view  of  the  abun- 
dant iodin  content  of  colloid  goiters.  Epidemics  of  goiter  presuma- 
bly are  the  results  of  an  infection  with  some  unknown  organism,  and 
possibly  the  endemic  form  has  a  similar  cause.  There  is  much  evi- 
dence, in  any  event,  that  whatever  the  cause  of  goiter  may  be,  it  often 
is  related  to  the  drinking  water,*'-  but  numerous  well-controlled  experi- 
ments fail  to  support  this  hypothesis."-''  Very  probably  the  causes 
of  colloid  goiter  and  parenchymatous  goiter  will  be  found  to  be  differ- 
ent from  the  causes  of  cystic  and  adenomatous  goiters. 

MYXEDEMA  AND   CRETINISM 

These  conditions  depend  upon  a  deficiency  of  thyroid  secretion, 
whether  from  operative  procedure  or  from  pathological  alterations  m 

59  See  Kooher,  ]\ritt.  a.  d.  Grenzfreb.  Med.  u.  Chir.,  1905.  vol.  14. 

coVirchow's  Aroli..   l!in-2    (16!)),  444. 

ei.Wien.  klin.  Woeli.,  1807    (10),  82.3. 

fi2  See  de  Quervain,  ]\Iitt.  a.  d.  Grenzgcb.  ^Icd.  n.  Chir.,  IDOo  (lo),  207 :  Birchcr, 
Zeit.  exp.  Path.  u.  Ther.,  1911    (9),  1. 

62a  See  Munch,  med.  Woeh.,  191.3  (60),  393  and  1813:  Sitzber.  ^Yion.  Akad.,  1914 
(123),  35. 


602  CHEMICAL    PATHOLOGY    OF    THE    DUCTLESS    GLAXDS 

the  oro-an.  ConsequtMitly  wo  find  evidences  of  a  decreased  protein 
metabolism,  the  urine  containing  a  diminished  quantity  of  nitrogen, 
especially  in  the  form  of  urea,  while  ammonia  and  other  forms  of 
nitrogen  are  relatively  excessive.  A  retention  of  nitrogen  and  phos- 
phorus has  been  found,  but  not  of  calcium  and  chlorine."^  The 
temperature  is  usually  subnormal,  and  the  energy  metabolism  is  low.*'^'^ 
Basal  metabolism  is  lower  than  in  any  other  known  condition  (Du 
Bois).*'^'^  Fat  and  carbohydrate  metabolism  seem  not  to  be  propor- 
tionately affected,"^  and  hence  the  elimination  of  COj  is  relatively  high 
as  compared  to  the  nitrogen  elimination.  Gastro-intestinal  disturb- 
ances are  common,  with  resulting  increase  in  the  amount  of  indican 
and  ethereal  sulphates  in  the  urine.  Whether  from  this  cause  or 
from  deep-seated  metabolic  anomalies,  there  is  a  decided  anemia,  and 
the  ability  of  the  corpuscles  to  combine  with  oxygen  seems  to  be 
decreased,  so  that  the  arterial  blood  may  contain  less  oxygen  than 
normal  venous  blood.  It  is  impossible  to  say  whether  the  failure  of 
growth  and  development  of  the  young  (cretinism),  and  the  mental 
and  physical  torpidity  of  the  adult,  are  due  to  an  autointoxication  from 
products  of  intermediary  metabolism  which  accumulate  because  of  the 
failure  of  the  thyroid  to  furnish  the  "stimulus"  necessary  for  their 
complete  destruction,  or  to  a  lack  of  some  essential  action  of  the  thy- 
roid secretion  upon  the  nervous  tissues  and  the  growing  cells  them- 
selves. Administration  of  thyroid  extract  to  cretinoid  children  causes 
retention  of  nitrogen  and  pliosphorus,  but  more  strikingly  of  cal- 
cium,*''' and  obese  cretins  lose  weight,  chiefly  from  the  non-nitrogenous 
elements  (Scholz).  The  amount  of  iodin  in  human  cretin  thyroids 
seems  not  to  have  been  estimated,  but  in  five  cretin  dogs  ^Marine  and 
Lenhart  could  find  no  thyroid  iodin  at  all. 

The  myxedematous  change  in  the  connective  tissues  is  in  the  nature 
of  a  reversion  to  the  fetal  type  of  tissue,  and  suggests  that  the  thyroid 
secretion  is  necessary  for  proper  cell  growth.  This  effect  might  be 
either  specific,  or  depend  simply  on  the  effect  on  protein  metabolism. 
Horsley'^"  describes  the  appearance  of  the  tissues  of  animals  dying 
after  thyroidectomy  as  follows :  ' '  The  subcutaneous  connective  tissue 
is  swollen,  jelly-like,  bright  and  shining,  and  excessively  stickA'.  The 
same  thing  is  observed  in  the  loose  tissue  of  the  mediastinum,  about 
the  heart,  and  in  the  omentum.  The  submaxillary  and  parotid  glands 
are  greatly  enlarged,   and  have  a  semi-translucent,   swollen   appear- 

"S  Benjamin  and  Rpuss,  Jalirl).  f.  Kinderlicilk.,  lOOS  (67),  201.  Tn  a  cretin 
Greenwaid  found  littlo  deviation  from  normal.  (Areli.  Tnt.  ^Vfed.,  1!)14  (14), 
374.) 

03aTalhot,  Aiiier.  .rour.  Dis.  Ciiil.,  inUi   (12),  1-1."). 

«3bArch.  Int.  Med.,  lOlO    ( 17) ,  nif). 

^14  Rarely  myxedema  and  diabetes  have  been  observed  cuiijoiiith'  (see  Strasser, 
Jour.  Amer.  ^ie<l.  Aasoe.,  1006   (44),  7f).'i). 

<i.-.  See  Iloiipardy  and  Lancstein.  Zeit.  f.  Kiiiderlieilk..  inO.'.  (til),  fi;?.'?.  Full 
fipures  are  {riven  bv  Seliolz,  Zeit.  exp.  Palli.  u.  Tlier,,  Iddd   (2),  270. 

""Brit.  Med.  Jo'ur.,  188.5    (i),  211. 


]i)\i:in:\i\  AM)  r/.7;77\ /M/ 


603 


anee;  from  the  cut  .surfae*e  a  sticky,  ylaiiy  fiuid  exudes.  Apparently 
the  parotid  becomes  transformed  into  a  mucous  gland ;  likewise  the  mu- 
cous membrane  of  the  alimentary  tract  is  swollen  and  transparent." 
Fetal  tissues  contain  normally  more  mucin  than  those  of  adults  (0.76 
per  cent,  as  against  0.37  per  cent,  in  the  subcutaneous  tissues,  accord- 
ing to  Halliburton),  and  in  the  early  stages  of  the  formation  of  ex- 
cessive subcutaneous  tissue  in  myxedema  such  an  increase  of  mucin 
may  be  present.  But,  under  ordinary  conditions,  the  term  myxedema 
seems  to  be  entirely  a  misnomer,  for  Halliburton's  analyses  showed 
that  the  skin  of  myxedematous  patients  contains  (juite  the  same  amount 
of  mucin  as  is  present  in  normal  skin."^  When  the  condition  is  of 
long  standing,  the  amount  of  mucin  may  even  be  much  reduced,  be- 
cause of  the  development  of  a  fibroid  character  in  the  connective  tissue. 
However,  in  monkeys  upon  which  thyroidectomy  had  been  perfonned, 
Halliburton  "**  found  a  decided  increase  in  the  mucin  in  the  tissues 
throughout  the  body,  especially  in  the  salivary  glands,  but  also  in  the 
skin,  subcutaneous  tissues,  and  tendons ;  and  mucin  was  found  in  the 
blood,  as  shown  by  the  following  table : 


Skin  and 

subcutane- 

Tendon 

Muscle 

Parotid 

Submax- 

Blood 

ous  tissue 

Normal  monkey   . 

0.89 

0.39 

0 

0 

a                  a 

0.9 

0.5 

0 

0 

0.1 

0 

After  thyroidectomy — 

55  days      .... 

3.12 

2.55 

0 

0.72 

0.0 

0.35 

33  days      .... 

trace 

49  days       .... 

2.3 

2.4 

trace 

1.7 

3.3 

O.S 

7   days        .... 

0.45 

0.904 

0 

trace 

0.16     1 

trace 

It  has  been  suggested  that  the  thyroid  produces  an  enzyme  which 
destroys  mucin,  but  that  such  is  the  case  has  never  been  demon- 
strated.*'^'^  Levin  "^  states  that  mucin  is  toxic  for  thyroidectomized 
rabbits,  but  this  is  not  substantiated  by  Nefedietf."'^ 

That  the  thyroid  is  connected  with  general  growth  is  shown  not 
only  by  the  thyroid  abnormalities  present  in  cretinism,  but  also  by 
the  frequent  observation  of  thyroid  defects  in  conditions  of  delayed 
growth  and  development  of  less  extreme  degree  {infantilism),  and  the 
favorable  eifeets  of  thyroid  feeding  in  many  such  cases.  Also  in  cer- 
tain types  of  short-limbed  dwarfs  {chondrodijsirophia  fatalis)  some 
thyroid  anomaly  may  have  an  etiologic  bearing,  for  in  such  a  case,  in 

•■w  Jour,  of  Patliol.  and  Bact.,   18!)3    ( 1 ) ,  90. 

08  Quoted  by  Horsley,  loc.  cit.  Later  experimenters,  however,  liave  had  difli- 
culty  in  producing  experimental  myxedema  as  described  by  Horsley,  or  have 
failed  entirely. 

Gsa  See  Parhon,  Compt.  Rend.  Soc.  Biol.,  1910   (79).  504. 

09  Med.    Record,    1900    (57),    184. 
ToVratch,  1901    (22),  Oct.  27. 


604  CHEMICAL    I'ATIIOLOGY    OF    THE    DUCTLESS    GLAyOS 

■\\iiicli  The  thyroid  was  liistolo^ically  greatly  altered  and  quite  free 
from  colloid.  I  could  find  no  trace  of  iodin.'^  On  the  other  hand,  the 
thyroid  of  a  giant  which  I  liave  analyzed  contained  62.9  mg.  of  iodin, 
or  six  times  the  amount  present  in  normal  glands. '- 

EXOPHTHALMIC  GOITER 

It  lias  by  no  means  been  conclusively  determined  that  exophthalmic 
goiter  is  due  to  an  intoxication  with  excessive  amounts  of  thyroid  se- 
cretion, either  normal  or  abnormal,  but  there  is  abundant  evidence  in 
favor  of  this  view.  IMost  important  is  the  similarity  of  exophthalmic 
goiter  to  the  effects  of  "hyperthyroidism"  or  " thyroidismus, "  pro- 
duced either  experimentally  or  through  overuse  of  thyroid  extract  for 
therapeutic  purposes.  In  thyroidisnuis  there  are  observed  a  rapid, 
Aveak  pulse ;  greatly  increased  metabolism,  especially  of  proteins ;  ^^ 
increased  secretion,  especially  of  perspiration;  marked  nervousness 
and  irrital)ility,  often  with  mental  confusion  and  delusions;  gastro- 
intestinal disturbances,  especially  diarrhea;  sweating,  flushing,  trem- 
ors, palpitation  of  the  heart,  loss  of  weight,  and  slightly  increased 
temperature  are  also  often  observed,  and  not  rarely  typical  exoph- 
thalmos may  appear."^^  These  manifestations,  which  are  common  to 
both  thyroidism  and  to  exophthalmic  goiter,  are  quite  the  opposite  of 
the  characteristic  changes  of  myxedema,  with  its  general  lowering  of 
all  metabolic  and  nervous  processes.  Reid  Hunt 's  acetonitrile  test  for 
thyroid  secretion  has  been  found  positive  in  the  blood  from  patients 
with  exophthalmic  goiter,'*  which  presumably  means  the  presence  of 
an  excess  of  thyroid  secretion  circulating  in  the  blood  in  this  disease. 
Furthermore,  the  histological  changes  observed  in  the  thyroid  may  re- 
semble those  of  compensatory  hypertrophy,  suggesting  strongly  that 
the  goitrous  change  of  this  disease  is  due  to  a  true  hypertrophy,  with 
increased  production  of  the  specific  secretions.  There  is  a  marked  in- 
crease in  the  mitochondria  of  the  thyroid  epithelium  in  exophthalmic 
goiter,  which  also  is  evidence  of  heightened  activity.'*-''  Kocher  '•'  says 
that  when  iodin  is  given  to  patients  with  cancer  of  the  thyroid  they 
may  develop  symptoms  of  exophthalmic  goiter,  as  if  an  excess  of  tliyro- 
iodin  were  formed.  Also  speaking  strongly  in  favor  of  the  view  that 
exophthalmic  goiter  is  the  result  of  overactivity  of  the  thyroid,  is  the 
frequent  cure  of  the  disease  through  removal  of  a  large  part  of  the 
diseased  gland.     Although  at  times  the  colloid  type  of  gland  is  found 

Ti  Reported   by   Hektoen,   Amer.   Jour.    :\I("(1.    Sci.,    1003    (125),   7.>1. 

"2  Reported  by  Bassoe,  Trans.  Cliicufio  I'atli.  Soe.,  1!)()3   (;1),  2'M. 

"•■''Metabolism  in  exopiitlialniic  j^oiter,  see  Du  ]?ois,  Areli.  Int.  Mi'd.,  l!'l(i  (17), 
*»l;i:    llalverson,   l?er-,reini   and    Hawk,   ihid.,   1010    (IS),  800. 

T.jii  Sufrar  niilizat ion  is  decreased,  as  sliown  by  study  of  tlie  utiii/ation  of 
suffar  {liven  intravenously   (\^'ilder  and  Sansuni,  Areh.  Int.  'Nted.,  1017   (10),  311). 

T4See  Gliedini,  Wien."  klin.  Woeli.,  1011  (-24),  73lJ:  Hunt  and  Seidell,  .lour. 
Pharm.  and  Exp.  Tlier.,  1010  (2),  1.5. 

7-«aCoet8eh,  Bull.  Johns  ITojjkins  Tlosp.,  101(1    (27),  120. 

■!^  Dent.  Zeit.  Chir..  1008    (01),  .302. 


j:\<)i'iiTii\i.\iic  (:<)iTi:i{  OOf) 

ill  ('.\()|)litlialiiii('  ^'•oitcr,  Mai'ine  contends  that  it  has  hccii  pn-ccdcd 
by  a  hyi)('i'j)histi('  stajiO."'' 

Based  on  tlie  tlieory  that  the  iioi-mal  function  of  tlie  thyroid  is  the 
detoxieation  of  metabolic  products,  is  the  sernni  treatment  advocated 
first  by  Ballet  and  Kni-i(iu('/.,  and  later  by  Lanz,  and  Burghart  and 
]-}lnnienthal;''  On  tlic  ])rinei})le  that  after  thyroidectomy  tiie  l)lood 
should  contain  an  acennudation  of  those  substances,  which  the  thyroid 
normally  neutralizes,  they  injected  the  serum  of  thyroidectomized 
goats  into  patients  with  exophthalmic  goiter,  in  the  hope  that  these  ac- 
cumulated  substances  might  in  turn  neutralize  any  excessive  thyroid 
secretion.  Favorable  results  were  obtained,  and  it  was  subsequently 
found  that  the  milk  of  thyroidectomized  goats  possesses  the  same  qual- 
ities, and  may  be  administered  by  mouth ;  this  has  led  to  ([uite  exten- 
sive clinical  use  of  this  method  of  treatment,  which  has  failed  to  show 
any  regidar  beneficial  effects  in  the  hands  of  most  careful  observers."-'' 
Of  similar  significance  are  the  favorable  effects  obtained  by  Beebe''* 
and  Rogers  '"  with  a  serum  made  by  immunization  of  animals  with  the 
mu'leoproteins  of  the  thyroid,  which  have  not  been  corroborated  by 
others. 

Oswald  *'*  found  that  the  thyroid  in  exophthalmic  goiter  contains 
generally  a  smaller  proportion  of  iodin  than  normal  glands,  but  with 
the  total  amount  approximately  normal.  However,  the  findings  are 
very  inconstant,  corresponding  with  the  fact  that  in  some  cases  of 
exophthalmic  goiter  the  amount  of  colloid  is  abundant  (in  which  case 
the  amount  of  iodin  may  be  large),  while  usually  the  amount  of  colloid 
is  small,  and  its  highly  vacuolated  condition  in  hardened  sections 
suggests  that  it  is  of  unusually  fluid  consistency.  A.  Kocher  *^  found 
that  either  the  amount  of  iodin  is  small,  which  is  usual,  or  else  very 
high,  but  it  is  seldom  the  same  as  in  normal  thyroids ;  the  more  dense 
the  colloid  in  the  follicles  the  higher  iodin  content  he  observed;  the 
phosphorus  content  is  both  relatively  and  absolutely  increased.  ^la- 
rine  has  found  that  in  exophthalmic  goiter  as  well  as  in  other  conditions 
the  amount  of  iodin  is  in  direct  proportion  to  the  colloid  and  inverse 
to  the  hyperplasia.  E.  V.  Smith  ^^''  obtained  in  simple  hyperplastic 
glands  an  average  of  0.54  mg.  of  iodin  per  gram  dry  weight,  as  com- 
pared with  1.52  mg.  in  hyperi)lastic  glands  showing  retrogressive 
changes  with  more  densely  staining  colloid.  P^onio  found  that,  as  with 
normal  thyroids,  the  physiological  effect  of  exophthalmic  goiter  glands 

70  See  also  Wilson.  Amer.  Jour.  Med.  Sci.,  lilOS    (136).  .S.jI. 

TTDeut.  med.  Woch.,  1899  (25),  627.  Also  Mobiua,  Miiiicli.  mo.l.  Woi-li..  I'.tOl 
(48),  185.3;  v.  Levden,  :\Ied.  Klinik.  1904  (1),  1;  Kuloiihor'r.  I'.crl.  kiiii.  Woch.. 
1905    (42),   3. 

"a  See  Sonne,  Zeit.  klin.  Mod..  1914    (?0),  229. 

78  Jour.  Amer.  ^Med.  Assoc,  1906   (4()),  484;   1900    (47).  6.)5. 

-^  Ihid.,  1900   (40),  487;   1006   (47),  001. 

soVirchow's  Areli.,   1902    (109),  475. 

siVirchow's  Ardi.,  1912    (208).  80. 

81a  Jour.  Amer.  Med.  Assoc.,   1914    i02).   113. 


606  CIIKMJCAJ,    PATflOLOaV    OF    THE    DVCTLEfi^    GLAyOS 

varies  directly  with  the  proijortioii  of  iodin,  and  such  glands  take  up 
iodin  administered  tlierajjeutically  just  as  a  normal  thyroid  does 
(Koeher,**-  i\Iarine  and  Leiiliart).^'^  These  results,  therefore,  indicate 
n()thiii<>-  either  for  or  a<i'ainst  the  liypothesis  that  exophtlialmie  <>-oiter 
is  due  to  autointoxication  with  the  secretion  of  the  thyroid,  but  Wilson 
and  Kendall  ^^^  find  that  in  the  toxic  type  of  goiters  there  is  but 
Vio-Vi  r.  as  much  of  the  active  iodin  compound  of  Kendall  as  in  normal 
glands,  and  hence  they  suggest  that  in  thyroid  intoxication  this  toxic 
material  has  been  discharged  from  the  thyroid  into  the  circulation. 

On  the  other  hand,  it  is  impossible  to  produce  a  symptom-complex 
resembling  exophthalmic  goiter  ®*^  in  animals  by  excessive  feeding  of 
thyroid,^*  either  normal  or  from  exophthalmic  goiter;  and  after  ex- 
tensive study  of  the  subject  Marine  and  Lenhart  have  come  to  the  con- 
clusion that  "the  essential  physiological  disturbance  of  the  thyroid  in 
exophthalmic  goiter  is  insufficiency,  its  reaction  compensatory  and  its 
significance  symptomatic."  This  view,  however,  certainly  fails  to 
agree  with  the  excellent  results  which  come  from  partial  extirpation 
of  the  thyroid  in  exophthalmic  goiter.  Oswald,-^  also  an  experienced 
investigator  in  this  field,  invokes  an  abnormally  irritable  nervous 
system,  which  stimulates  the  thyroid  and  in  turn  is  stimulated  by 
the  thyroid  secretion,  constituting  a  vicious  circle.  Other  observers 
are  of  the  opinion  that  not  an  excessive,  but  a  perverted,  secretion  is  at 
fault,^**  a  view  not  confirmed  by  tests  of  the  effects  of  thyroid  extracts 
on  animals.^^  However,  it  is  stated  by  Blackford  and  Sanford,*^  that 
extracts  of  the  thyroid  in  this  disease,  as  well  as  the  blood  of  patients 
in  the  acute  toxic  stages,  exhibit  a  marked  depressor  effect  on  blood 
pressure,  which  is  distinct  from  that  of  choline,  and  which  they  believe 
to  be  specific  for  exophthalmic  goiter. 

There  can  be  no  doubt  that  the  thyroid  secretion  is  capable  of  caus- 
ing serious  intoxication,  for  patients  who  have  overused  thyroid  prep- 
arations in  the  treatment  of  obesity,  skin  diseases,  etc.,  have  often  suf- 
fered severely  from  the  symptoms  mentioned  previously,  and,  in  at 
least  one  such  case,  a  diagnosis  of  exophthalmic  goiter  was  made  be- 
fore the  cause  of  the  disturbance  was  detected.  Not  infre(|uently 
evidences  of  acute  intoxication  have  followed  immediately  after  opera- 
tions upon  the  thyroid,  and  these  have  been  considered  as  due  to 
intoxication  with  the  large  quantities  of  thyroid  secretion  that  have 

R2Arcli.  klin.   Cliir..   1010    (1)2).  442;    1!)11    (90),  403. 

83Arfli.  Int.  Mi'd.,   1!»11    (S),  205. 

!^3a  Anier.  Jour.  Mod.  Sci.,   1010    (151),  70. 

'<4aT]if>  patlio<roni'sis  of  tlie  exoplitlialmos  is  iiiikiiowii.  Seo  Trocll,  .Vicli.  Int. 
Mod.,  1016    (17),  382. 

«t  See  Carlson  et  al.,  Anwr.  .lour.  I'lnsioi.,  1012  CM)).  120;  Marino,  .lour.  Anicr. 
Med.  Assoc,  1012   (50),  325. 

t*"' Correspdiidcnzlilatt  Scliweizor  Aerzto,  1012    (42).  1130. 

»"Klose  rt  al.,  Hcitr.  z.  klin.  Cliir..  1012    (77),  001. 

><"  See  Sclir.nltorn,  Arch.  exp.  I'atli.  u.  riiarni..   1000    i(;o),  300. 

f'WJour.  Anier.  Med.  Assoc.,   1014    (02),   117. 


EXoi'iiriiM.Mic  (;<)iii:i{  607 

escaped  from  llic  <:l;iii(l  (lui-iiiii-  the  njx'i-at  i\r  iii;iiii|Mil;it  ion.  Tlic  fact 
tliat  (i))ibli/(jpi(i,  r('S('iiil)liii<z-  that  prodiK-ctl  hy  tol)acc(),  etc.,  may  follow 
overuse  of  thyroid  preparations  ''■'  is  also  indicative  of  their  toxicity,  as 
also  is  the  glycosuria  that  may  i-esult  from  thyroid  administration.*"'' 

Even  if  the  hypotlie.sis  that  ex()])hthalmic  goiter  is  due  to  intoxi- 
cation wilh  thyroid  secretion  is  correct,  we  have  no  satisfactory  ex- 
planation of  the  cause  of  tlie  hyperactivity  of  the  thyroid.  In  some 
cases  deji'enerative  changes  have  been  observed  in  the  superior  cervical 
sympathetic  ganglia,  and  cure  or  improvement  of  exophthalmic  goiter 
is  said  to  have  followed  resection  of  these  ganglia;  however,  this  re- 
lation has  not  been  observed  at  all  constantly.  In  other  cases  there 
has  been  evidence  that  suggested  a  primary  intoxication  with  the  i)rod- 
ucts  of  intestinal  putrefaction,  leading  to  a  secondary  hyperplasia  of 
the  thyroid,  but  this  also  seems  to  be  an  exceptional  observation.  All 
things  considered,  it  seems  most  probable  that  the  hyperactivity  of 
the  thyroid  is  due  to  some  exciting  condition,  and  is  not  of  itself 
primar3%  although  the  resulting  hypersecretion  of  the  thyroid  may 
cause  the  dominant  features  of  the  disease.  The  frequent  association 
of  exophthalmic  goiter  with  puberty  and  pregnancy  suggests  that  some 
abnormality  in  the  function  of  the  generative  organs  may  be  a  frequent 
starting-point  of  the  thyroid  derangement.-''"''  In  not  a  few  cases  dia- 
betes or  pancreatitis  have  been  associated,""  and  some  observers  state 
that  the  pressor  substance  (presumably  epinephrin)  in  the  blood  is 
much  increased  in  exophthalmic  goiter.''^ 

The  Relation  of  the  Parathyroids  to  Exophthalmic  Goiter. — 
This  has  not  yet  been  definitely  established.  As  nervous  manifesta- 
tions are  very  prominent  after  parathyroidectomy,  so  that  many  ex- 
perimenters attribute  all  the  acute  nervous  and  muscular  symptoms 
of  total  thyroidecton\y  to  simultaneous  removal  of  the  parathyroids, 
it  has  seemed  very  probable  that  these  organs  may  be  more  closely  asso- 
ciated with  exophthalmic  goiter  than  is  the  thyroid  itself."-  Against 
the  hypothesis  that  exophthalmic  goiter  is  due  to  parathyroid  insuf- 
ficiency, however,  stand  the  following  facts : 

(1)  Kemoval  of  one  lobe  of  the  thyroid  often  causes  improvement 
or  recovery  in  this  disease,  yet  with  the  lobe  of  the  thyroid  is  gener- 
ally removed  the  adjacent  parathyroid,  which  would  decrease  the 
amount  of  parathyroid  tissue,  and  make  worse  any  existing  parathy- 
roid   insufficiency.      (2)    Therapeutic    administration    of    parathyroid 

S9  Birch-Hirsclifeld  and  Inouve,  Graofe's  Aroli..   inOo    (til),  4nn. 

89b  See  Geyelin,  Arch.  Int.  Med.,  1915  ( 16),  97o. 

S9a  The  serum  of  patients  with  exophthalmic  ofoiter  sliows  by  Ahdcrhalden's 
method  a  constant  power  to  digest  tliyroid  tissue,  and  sometimes  ovarv  or  testich^ 
(Lampe  and  Fuchs.  Miihch.  med.  Wo'ch.,  1913   (BO),  No.  39). 

90  Thompson,  Amer.  -lour.  Med.  Sci.,  1906  (132),  835;  Cohn  and  Peiser.  Deut. 
med.  Woch.,   1912    (3S),  60. 

91  Brokinjr  and  Trcndelonbiirnr.  Dent.  Arch.  klin.  Med.,   1911    (103),   168. 

92  This  subject  is  tliorouphlv  reviewed  by  MacCallum,  Med.  News,  1903  (83), 
820;  Iversen,'Arch.  Internat.  de  C'hir.,  1914    (6),  255. 


608  CHEMICAL    PATHOLOdY    OF    THE    DUCTLESS    GLAXDS 

tissue  or  extract  has  had  no  significant  effect  on  the  disease.  (3)  Xo 
considerable  or  characteristic  anatomical  changes  occur  in  the  para.- 
thyroids  in  exophthalmic  goiter,"^  while  the  great  majorit}'  of  all  cases 
show  changes  in  the  thyroid.  (4)  Tlic  parathyroids  seem  to  have  but 
slight  influence  on  metabolism  (MacC'allum),  while  metabolic  abnor- 
malities are  very  marked  in  exopiithalmie  goiter.'-'"' 

THE  ADRENALS  AND  ADDISONS  DISEASE  03 

Like  the  hypophysis,  the  adrenals  are  essentially  double  organs, 
containing  nervous  and  glandular  tissues.  The  medulla  is  of  sym- 
pathetic nervous  system  origin,  a  part  of  the  chromaffin  system, 
which  in  mo'st  animals  is  enclosed  in  a  layer  of  entirely  different  na- 
ture and  origin :  the  cortex  being  an  epithelial  structure,  derived  from 
the  urogenital  anlage,  and  resembling  niost  closely  in  structure  (and 
perhaps  in  function)  the  corpus  luteum  of  the  ovary.  In  some  marine 
animals,  indeed  (eels,  sharks,  etc.),  the  sympathetic  tissue  portion  and 
the  cortical  tissue  exist  as  separate  organs. 

The  adrenal  cortex  seems  to  be  related  especially  to  the  generative 
system,'-"*  as  shown  by  the  following  facts : 

1.  The  embryologic  origin  in  the  urogenital  anlage,  and  the  histo- 
logic structure  which  is  similar  to  the  corpus  luteum. 

2.  In  many  animals  there  occurs  hypertrophy  of  the  cortex  during 
the  breeding  season,  and  there  are  histological  ditt'erences  in  the  glands 
of  males  and  females  (Kolmer). 

3.  Many  cases  of  sexual  precocity  have  been  observed  in  association 
with  tumors  or  hypertrophy  of  the  adrenal  cortex ;  and  defective  sex- 
ual development  has  been  found  associated  with  atrophj^  of  this  tis- 
sue.^^ 

4.  The  medulla  increases  relatively  little  in  size  after  birth,  while 
the  cortex  increases  with  the  development  of  the  individual. 

Whether  the  cortex  has  other  functions  or  not  is  not  yet  known."''' 
Biedl  has  found  evidence  that  cortical  substance  is  essential  for  life.'''*' 
Animals  with  accessory  adrenals,  which  contain  only  cortical  substance, 
withstand  ablation  of  the  adrenals  proper,  presumably  because  the  rest 
of  the  chromaffin  substance  remains  to  compensate.""''  Chemically  the 
cortex  is  characterizetl  by  not  containing  the  specific  vaso-constrictor 

»3  MacCallum,  JoJms  Hopkins  ITosp.  T^ull..   1005    (Ifl),  2S7. 

94  The  calcium  excretion  in  exophtlialmic  >;oit('r  ])aralh'ls  tlic  nitro^icn  (Towles. 
Amer.  Jour.  Med.  Sci.,  1!)10    (140),   100). 

"•'■'Literature  <fiven  l)v  Uaver,  Kr^^ehnisse  Patliol.,  1!U()   (XIN'.t,  1. 

!>'■•  See  Kolmer,  lMlfi->er"s  .\rcli.,  ]!H2    (144),  .'Itil. 

97  See  (JIvnii,  Quart,  .lour.  Med.,  litl'i  I.")),  157;  .lump  ct  al..  Amer.  .lour.  Med. 
Sci.,  1014    (147),  5(iS. 

f»7a  It  does  not  liave  a  marked  ell'ect  on  tlie  development  of  tadpoles,  lience  dif- 
ferinj,'  from  thyroid  and  tiiymus    ( (Judernatseli ) . 

'J"i>  See  also  Crowe  and  \\"ish)cki.  Bull,  .lohns  Hopkins  llosp.,  1!)14    ('25).  287. 

"7c  See  Fulk  and  MacLeod  (Amer.  .liMir.  i'iiysiol.,  lilHl  (40),  21)  wlio  found 
that  the  active  j)rinci])le  of  otlier  cliromallin  tissues  lias  tlie  same  physiok)fricaI 
efTect  as  tliat  of  the  adrenal   medulla. 


THE  ADh'i:\.\f.S    \\l>    \ltn/sn\-s  DISEASE  609 

Iji-iiiciplc.  lilt'  cpiiu'phi'in,  and  by  containing  a  very  large  proportion  of 
lil)()i(ls.  Thus,  in  water-free  human  adrenals  (cortex  and  medulla 
botli  iiu'ludcd)  there  was  found  36.;^  per  cent,  of  ether-soluble  material, 
of  wliich  20. ()  per  cent,  was  cholesterol  and  33  per  cent,  was  let-ithin.'-"* 
The  proportion  of  fats  and  lipoids  varies  greatly  during  changes  of 
age,  disease,  and  perhaps  of  function,  and  there  are  those  who  believe 
the  adrenal  cortex  to  be  a  chief  source  of  the  lipoids  of  the  blood,  to 
which  much  imi)ortant  function  is  ascribed  in  the  reactions  of  immu- 
nity. (See  Lipoids,  under  Fatty  ^[ctamorpliosis.)  When  cholesterol 
is  fed  in  large  amounts  some  is  deposited  in  the  adrenal  cortex,"'-'  while 
in  nuiny  diseases,  notably  delirium  tremens  ( Ilirsch ) ,"'-"'  the  lipoid 
content  of  the  adrenals  is  greatly  decreased.  In  renal  and  arterial 
disease  the  adrenal  lii)oids  have  been  found  increased.''"*^  The  lipins 
of  the  adrenal  cortex  are  said  to  contain  little  or  no  neutral  fat,^"*"  but 
free  fatty  acids  which  may  be  increased  when  the  cholesterol  decreases. 
Loss  of  body  fats  is  not  accompanied  by  a  loss  of  adrenal  lipoids 
ordinarily,  although  they  decrease  in  acute  infections,  especially  pneu- 
monia.'""^ A  vaso-depressor  effect  is  produced  by  extracts  of  adrenal 
cortex,  probably  caused  by  choline  which  has  been  found  in  such 
extracts. 

The  medulla  is  characterized,  besides,  by  its  pigmentary  content, 
by  the  remarkably  active  internal  secretion,  epinephrin,^  which  it 
always  contains  in  greater  or  less  amount.  Presumably  epinephrin, 
of  which  the  formula  is 

HO  /^"^CIIOH  — CH,   (XH)   —  CH3 
HO 

is  derived  from  the  aromatic  radical  of  the  proteins,  its  close  relation- 
ship to  tyrosine  being  seen  when  the  formula  of  the  latter  is  com- 
pared 

ho/      \cH„  — CH    (XH,)   —  COOH 

That  epinephrin  is  formed  from  tyrosine  directly,  is,  however,  not 
yet  demonstrated.  There  are  also  other  amines  and  aromatic  com- 
pounds which  might  be  formed  in  the  body,  that  have  a  pressor  effect, 
and  which  perhaps  are  formed,  although  not  yet  identified.-     It  is  to 

98  Wells,  Jour.  INIed.  Res.,  1!)08    (17),  461. 
99Krylov,  Beitr.  patli.  Anat.,  Iftl4    (.icS),  434. 
»9a  Jour.  Amer.  :\led.  AssOc,  ini4    (63),  2186. 
99bChauffard,  Compt.  ?>end.  Sof.  Biol..  1914    (76),  529. 
99c  FJorberg.  Skand.  Areli.  Plivsiol.,  191")    (32),  2S7. 
99d  Elliott,  Quart.  Jour.  Mcd.^  1914    (S).  47. 

1  Tins  name,  given  by  Abel  and  Crawford,  is  to  be  preferred  to  the  others  in 
common  use,  especially  the  niost-iised  term  "adrenalin,"  wliich  has  been  copy- 
riplited  bv  a  manufacturinif  establishment  so  that  this  name  means  specifically 
tiieir  product,  and  not  the  active  principle  of  tlie  adrenal  from  whatever  source. 

2  See  Barjrcr  and  Dale,  Jour.  Plivsiol..  1910    (41),  19. 

39 


610  CHEMICAL    PATHOLOGY    OF    THE    DUCTLESS    GLAXDS 

be  borne  in  mind  that  the  formation  of  ei)inep]irin  is  not  limited  to  tbe 
adrenals,  but  that  otlier  islands  of  chromaffin  sympathetic  tissue  can 
do  the  same,^  which  explains  the  observed  discrepancies  between  the 
anatomic  chantres  in  the  adrenals  and  the  clinical  manifestations  of  a 
deficiency  in  epinephrin. 

Accordino-  to  Goldzieher  ^  the  normal  human  adrenals  contain  to- 
gether about  4  mg.  epinephrin,  which  ma}'  be  increased  in  conditions 
with  high  blood  pressure,  such  as  arteriosclerosis  and  nephritis,  in 
which  he  found  an  average  of  5.8  mg. ;  and  in  septic  conditions  with 
low  pressure  he  found  it  reduced  to  an  average  of  1.5  mg.*^  The 
human  adrenal  contains  no  epinephrin  before  birth,"'  but  Fenger  ^ 
found  it  present  in  the  adrenal  of  unborn  domestic  animals.  Autolysis 
of  the  adrenal  decreases  the  amount,'  but  not  all  of  the  epinephrin  is 
destroyed  even  several  days  after  death,  as  shown  by  Ingier  and 
Schmorl,^  who,  using  both  morphological  and  chemical  methods,  also 
found  a  gradual  increase  in  the  epinephrin  content  of  normal  glands 
from  birth  to  the  ninth  year,  after  which  it  remains  practically  con- 
stant at  about  4.5  mg.  (males  4.4,  females  4.71  mg.).  They  also  found 
a  slight  increase  in  arteriosclerosis,  more  in  acute  and  chronic  nephritis, 
and  a  decrease  in  diabetes  and  narcosis,  there  being  practically  no 
epinephrin  in  the  adrenal  of  Addison's  disease.  In  most  of  the  infec- 
tious diseases  they  found  no  changes,  and  in  amyloid  infiltration  the 
amount  was  about  normal.  The  amount  of  chromaffin  substance  and 
epinephrin  do  not  always  run  parallel,  although  Borberg  "  found  a 
close  parallelism ;  this  author  also  failed  to  observe  any  marked  de- 
crease of  chromaffin  substance  in  narcosis.  Elliott ""''  found  a  low 
epinephrin  content  in  acute-infectious  diseases,  and  especially  low  in 
acute  cardiac  failure  associated  with  great  mental  distress:  he  did  not 
find  any  increase  in  the  epinephrin  in  nephritis  or  in  any  otlier  dis- 
ease. 

The  function  of  the  epinephrin  is  manifestly  to  modify  the  tone  of 
the  non-striated  muscle  fibers  which  are  under  control  of  the  sympa- 
thetic nervous  system,  acting  upon  some  receptive  substance  present 
in  the  muscle,  perhaps  at  the  nerve  endings.  But  it  is  a  fact  of  much 
practical  importance  that  administration  of  epinephrin  will  not  com- 
pensate successfully  for  the  loss  of  the  adrenals,  whether  because  the 
gland  secretes  other  things,  or  because  the  intermittent  artificial  ad- 
ministration of  the  epinephrin  will  not  compensate  for  the  regulated 

3  See  Vincent,  Proc.  Ro.y.  Soc.,  B,  1908   (82),  502. 

4Wion.  klin.  Wooh.,    1010    (23),   SOn. 

•1"  See  also  Keicli  and  Bcresne-j^owski,  T?eitr.  klin.  Chir..  1!)14  (91),  40.'^.  Oliiui 
(Verb.  .Japan.  Palli.  riescll.,  lOKJ  (('.).  l.'j)  found  tlie  normal  content  lo  ho  ahout 
5.0  mg.,  averai,nn<r  S.32  inj,'.  in  clironic  nei)liritis. 

■'■' Moore  and  Pnrinton,  Amor.  Jo;:r.  Plivsioi.,  1!)00  (4).  51  ;  .Fulian  Lewis  .Tour 
Biol.  Cliem..  1010  (24),  240. 

'i.Ioiir.   Biol,  ('hem.,   1012    (II),  480. 

7  C'omeHKJitti,  Areli.  exj).  Palli.  u.  Pliarm.,   1010    (02),  100. 

M)eut.  Aroh.  klin.  :Med..  1011    (104),   125. 

y  Skand.  Arcli.  Plivsiid,,   1012    (27),  341;   101.3    (28).  01. 


THE  Ainn:\.\i.s  A\D  .\nn/s()\-s  disease  611 

secivtion  of  the  ^laiul  uiidfi'  normal  conditions,  or  both.  It  would 
seem  tliat  tlie  adrenal  has  an  elt'ect  on  other  glands,  for  injections  of 
epinephrin  cause  glycosuria  in  animals,  as  also  does  manipulation  of 
the  adrenals,  or  i)aiiitiii^-  llie  epinephrin  on  the  pancreas.  There  is 
much  disagreement  as  to  tlie  effects  of  extirpation  of  the  adrenals  on 
carbohydrate  metabolism,  and  the  nature  and  cause  of  the  effects  ob- 
served. Jiiedl  sums  up  tlie  evidence  with  the  statement  that  the  in- 
ternal secretion  of  the  chromaffin  system  is  of  importance  in  the  mo- 
bilization of  the  sugar  of  the  blood,  and  the  formation  of  the  glycogen 
in  the  tissues.  That  the  adrenal  is  at  all  im})licated  in  human  diabetes 
has  not  been  demonstrated. 

Acute  insufficiency  of  the  adrenals,  caused  most  often  by  hemorrhagic 
infarction,  but  sometimes  by  other  lesions,  may  cause  sudden  collapse, 
asthenia  or  death.'"  The  extent  to  which  the  cortex  and  medulla  re- 
spectively are  responsible  is  undetermined.  The  French  authors  espe- 
cially lay  great  weight  on  adrenal  insufficiency  as  a  cause  of  patho- 
logical states. ^^  Surgical  shock  has  also  been  attributed,  at  least  in 
some  cases,  to  exhaustion  of  the  adrenals,  which  takes  place  under  the 
influence  of  the  anesthetic  and  the  stimulation  to  the  nervous  system 
by  the  operative  manipulation,  perhaps  augmented  hy  concurrent  in- 
fections.'- 

It  is  possible  that  in  some  cases  of  trauma  to  the  adrenal,  acute  hem- 
orrhage or  infection,  intoxication  from  an  excess  of  epinephrin  might 
occur,  but  it  is  improbable  that  fatal  results  could  be  produced  in  this 
way,  for  the  lethal  dose  for  dogs  and  rabbits  is  about  0.1  to  0.25  mg. 
per  kilo,  and  the  two  adrenals  in  man  contain  in  all  but  about  4  to  5 
mg.  epinephrin.  IModema,  how^ever,  states  that  there  is  so  much  epi- 
nephrin set  free  after  hemorrhage  into  the  adrenal,  that  it  can  be  dem- 
onstrated microchemically  in  the  liver,  and  that  the  symptoms  and 
autopsy  findings  are  identical  with  those  of  acute  epinephrin  intoxica- 
tion. In  animals,  repeated  doses  of  epinephrin  produce  a  decreasing 
effect,  not  only  on  blood  pressure  but  on  the  glycosuria  and  other  symp- 
toms, indicating  an  acquirement  of  tolerance,  but,  because  of  its  non- 
protein nature,  epinephrin  does  not  cause  the  production  of  anti- 
bodies.'^ 

IMany  studies  have  been  directed  to  determine  the  relation  of  the 
adrenal  to  hypertrophy  of  the  heart  and  to  interstitial  nephritis  with 
high  blood  pressure.  Some  have  found  more  or  less  increase  in  size  in 
the  adrenals  under  these  conditions,  chiefly  involving  the  cortex,  and  a 
slight  increase  in  the  epinephrin  content  has  been  reported,  but  it  is 

10  Literature  bv  Lavenson,  Areh.  Int.  ^Med.,  inOS  (2),  62;  Materna.  Ziejiler's 
Beitr.,  1910   (48)',  236. 

11  See  Sergent,  Presse  'SUd..  HlOO  (17),  480;  S^zary,  Semaine  :M^d..  1012 
(33),   61. 

12  See  Hornowski.  Arch.  nied.  exper..  l!)()0  (21),  702;  Virohow's  Arcliiv.,  100!) 
(108),  03. 

13  See  Elliott  and  Durham,  Jour,  of  Physiol.,   1006    (34),  430. 


612  CHEMICAL    PATHOLOGY    OF    THE    DUCTLESS    GLANDS 

veiy  doubtful  if  these  observations  are  of  significance."  It  has  been 
reported  by  several  investigators  that  the  blood  in  such  conditions  con- 
tains sufficient  epinephrin  to  permit  of  its  detection  and  measurement 
by  its  physiological  effects  (dilatation  of  the  frog's  iris,  contraction 
of  the  rabbit  uterus  or  blood  vessels,  inhibition  of  contraction  of  the 
intestine).  The  critique  of  this  work  by  Stewart,^'''  however,  makes 
it  necessary  to  discount  most  of  the  published  results,  as  being  in- 
adequately^ controlled.  He  found  no  epinephrin  even  in  blood  coming 
direct  from  the  adrenal  veins,  unless  the  gland  had  been  stimulated 
or  manipulated,  and  none  could  be  detected  in  the  serum  from  several 
patients  with  high  })ressure  from  various  causes,  as  well  as  in  mental 
disturbances  and  exophthalmic  goiter.  Vaso-constrictor  substances 
may  be  present  in  serum,  both  normal  and  pathological,  which  are  not 
epinephrin.  His  negative  results  are  corroborated  by  Janeway  and 
Park.^"  Broking  and  Trendelenburg,^^  using  a  perfusion  method 
which  the}^  believe  to  be  reliable,  found  a  normal  pressor  effect  from 
the  blood  of  persons  with  arteriosclerosis  and  high  blood  pressure,  a 
decrease  in  nephritis  with  high  pressure,  a  great  increase  in  exophthal- 
mic goiter,  and  no  changes  in  pregnancy,  chlorosis  and  diabetes. 

Arterial  Degeneration  from  Epinephrin. ^'^ — An  interesting  result 
of  repeated  injections  of  epinephrin  into  animals  is  the  appearance  of 
a  marked  atheromatous  degeneration  of  the  aorta,  with  calcification. 
This  was  first  observed  by  Josue,  and  since  confirmed  by  Erb,  Fischer, 
Gouget,  Loeb  and  Githens,  and  many  others.  These  lesions  are  quite 
different  from  those  of  human  aortic  arteriosclerosis,  the  chief  change 
being  degeneration  of  the  muscle-cells  of  the  media,  without  any  con- 
siderable inflammatory  reaction.  There  is,  however,  more  resemblance 
to  the  atheromatous  changes  seen  in  the  arteries  of  the  extremities. 
They  do  not  seem  to  be  due  to  the  heightened  blood  pressure,  since 
simultaneous  administration  of  substances  that  keep  the  blood  pressure 
down  does  not  prevent  the  atheroma  from  developing  (Braun),  while 
other  substances  that  raise  blood  pressure,  such  as  nicotine  (Josue) 
or  pyrocatechin  (Loeb  and  Githens),  do  not  cause  atheroma.  Pre- 
sumabl}',  therefore,  epinephrin  causes  the  arterial  changes  by  a  direct 
toxic  action,  but  the  influence  of  increased  blood  pressure  cannot  be 
entirely  excluded.  However,  slow  injection  of  epinephrin,  so  regu- 
lated that  there  is  an  increase  in  the  blood  content  without  significant 
rise  of  pressure,  fails  to  produce  arteriosclerosis.'"*"  Myocardial  de- 
generation is  also  observed  in  experimental  aninmls,  and  later  may  lead 
to  an  interstitial  myocarditis   (Pearce).     These  experiments  suggest 

14  See  Pearce,  .lour.  Kxpcr.  ^fcd.,  lOOS  (10),  73.'):  Tliomas,  Zie<iler's  Boitr., 
1910    (4!)),  228. 

15  Jour.  Exp.  Mod.,   IHll    (14),  .377:    1012    (IT)).  .147. 
I's.Tour.  Exp.  Med.,   1012    (16),  541. 

iTDcut.  Arc'li.  klin.  Med.,  1011    (103),  lOS. 
•    i«Li(erature  <riven  l.y  Saltykow,  ("eiil.   f.   Tatli..   lOOS    (10).  3()0. 
ixh  van  T-ecrsutii  and' Hassc'rs,  Zeit.  cxp.   Pa(l\..    1014    (Itt),  230. 


MHU SOX'S  DISEASE  613 

the  ))()ssil)ility  that  (ivci'sccrct  ion  of  ('])iii('|)hi'iii  may  be  a  cause  of 
artei'iosclcrosis,  I)iit  tlici'c  is  no  evidence  tliat  this  actually  occurs  in 
man. 

ADDISON'S  DISEASE  i  ' 

As  pointed  out  bel'oi'c,  the  ])i-ot'oun(l  deficiency  in  the  de])rossor  prin- 
ciples evident  in  the  manifestations  of  Addison's  disease  implies  loss 
of  function,  not  only  of  the  adrenal  medulla,  but  also  of  the  rest  of 
the  chromaffin  tissues  which  produce  this  same  sort  of  material. 
Therefore  it  is  possible  to  have  any  amount  of  destruction  of  the  ad- 
renals without  Addison 's  disease,  if  there  is  sufficient  compensation 
by  the  other  chromaffin  structures,  or,  conversely,  Addison's  disease 
may  occur  when  the  adrenals  seem  morphologically  little  altered,  which 
occurs  in  about  10  per  cent,  of  all  cases.  In  typical  cases,  however, 
the  adrenals  have  been  found  entirely  devoid  of  epinephrin,-"  and 
usually  the  structural  alterations  are  conspicuous.  While  some  have 
held  that  the  destruction  of  the  adrenal  cortex  is  of  importance  in  Ad- 
dison's  disease,  this  does  not  seem  to  have  been  conclusively  demon- 
strated. 

The  pifi-mentation  of  the  skin  -^  has  not  yet  been  explained,  but  in 
view  of  the  fact  that  oxidizing  enzymes  readily  convert  epinephrin, 
tyrosine,  and  related  aromatic  substances  into  pigments,  and  that  in 
Addison's  disease  we  have  a  deficiency  in  a  tissue  which  is  known  to  be 
concerned  in  the  metabolism  of  aromatic  compounds,  it  seems  probable 
that  the  pigmentation  is  the  result  of  this  defective  metabolism  of  the 
chromogenic  aromatic  compounds.  In  support  of  this  view  is  the  ob- 
servation of  Bittorf  -^^  that  the  skin  of  persons  with  Addison's  disease 
has  an  augmented  power  of  oxidizing  epinephrin  and  tyrosine  to  pig- 
mented substances.  Until  the  pigment  of  Addison's  disease  has  been 
isolated  and  analyzed,  however,  this  hypothesis  will  probably  remain 
an  hypothesis.  (See  pigmentation.  Chap,  xvi.)  Addison's  disease 
can  occur  without  pigmentation. 

That  there  is  a  deficiency  in  the  formation  of  epinephrin  is  at- 
tested by  the  low  blood  pressure  and  general  low  tone  of  the  unstri- 
ated  muscle  tissue.  Carbohydrate  metabolism  is  also  altered,  Porges  -- 
having  found  hypoglucemia  in  Addison's  disease,  and  an  increased 
sugar  tolerance  having  been  observed  by  others.  Whether  the  ad- 
renals exert  a  detoxicating  eflfect,  and  the  symptoms  of  the  disease 
are  partly  the  result  of  an  autointoxication  of  some  sort,  is  at  present 
unknown,  although  this  idea  has  often  been  advanced.     The  general 

19  LitfM-aturo  on  Clicinistrv,  bv  Eiselt,  Zeit.  klin.  ^[ed.,  1010   (60),  303. 

20  Inkier  and  Schniorl.  Dent.  Arcli.  klin.  Mefl.,  1011    (104),  12ri. 

21  Aooordin<r  to  Straiib  (Dent.  Arch.  klin.  'Mod.,  1000  (07).  67)  pigmentation 
may  occur  within  17  da^•a  after  thrombosis  of  tlie  adrenal  vein. 

2'iaArch.  exp.  Path.,  1014    (75),  143. 

22  Zeit.  klin.  :Med.,  1000  (60),  .341;  also  Bernstein,  "nerl.  klin.  Woch..  1011  (4S), 
1794.  Normal  blood  sugar  was  found  bv  Broekmever,  Deut.  nied.  ^N'och.,  1014 
(40),  1562. 


614  CHEMICAL    PATHOLOGY    OF    THE    DCCTLEHH    GLANDS 

metabolism  of  Addison's  disease  shows  no  very  striking  or  character- 
istic chang-es,  over  and  above  tliose  associated  with  the  emaciation. 
Wolf  and  Thacher  -^  found  a  decrease  in  endogenous  creatine  and 
purine  excretion,  and  some  evidences  of  acidosis  towards  the  end  of 
the  disease ;  deaminizing  power  and  oxidation  of  cystine  sulphur  to 
SO4  were  not  impaired.  Eiselt  believes  that  there  is  a  toxocogenic 
loss  of  tissue. 

Administration  of  adrenal  tissue  and  extracts,  or  epinephrin, 
whether  by  mouth  or  subcutaneously,  is  not  efifective  in  ameliorating 
the  course  of  Addison's  disease,  at  least  in  most  cases.  Thus,  in  97 
cases  collected  by  Adams, -^  adrenal  treatment  caused  some  improve- 
ment in  31,  43  M^ere  not  benefited,  7  were  made  M^orse,  while  16  were 
described  as  permanently  improved.  The  most  favorably-  affected 
is  usually  the  muscular  and  gastro-intestinal  asthenia,  while  the  pig- 
mentation is  not  usually  altered.  There  is  little  effect  on  metabo- 
lism.-'^ 

THE  HYPOPHYSIS  AND  ACROMEGALY  ai 

Although  the  hypophysis  contains  in  its  anterior  lobe  and  in  the 
pars  intermedia,  a  certain  number  of  spaces  filled  with  colloid  and  re- 
sembling the  alveoli  of  the  thyroid  in  appearance,  there  is  no  evidence 
that  an  appreciable  amount  of  iodin  is  present  here  except  when  thera- 
peutically administered.^-  The  posterior  lobe  contains  an  active  diu- 
retic and  pressor  substance,^*  the  exact  nature  of  which  is  not  yet 
known,  although  in  many  respects  its  action  resembles  that  of  epine- 
l)hrin.  It  seems  less  active  in  producing  arteriosclerosis  than  is  epine- 
phrin, and  its  pressor  effects  are  of  longer  duration.  It  seems  to 
stimulate  smooth  muscle  without  respect  to  innervation  (thus  differing 
from  epinephrin),  but  with  a  special  potency  in  stimulating  contrac- 
tions of  the  uterus ;  and  hence  it  has  a  wide  clinical  use  under  the  name 
pituitrin.  The  chemical  nature  of  pituitrin  is  not  yet  determined,  but 
it  seems  to  be  closely  related  to  ^-iminazolylethylamine,  the  pressor 
base  derived  from  histidine,  and  which  also  stinnilates  uterine  con- 
tractions. Injection  of  the  posterior  lobe  extract  lowers  the  assimila- 
tion limit  for  carbohydrates  and  causes  glycogenolysis  (' rushing")  .^^ 
and  is  a  powerful  galactagogue  (Ott). 

Removal  of  the  anterior  lobe  of  the  gland  in  young  animals  is  fol- 
lowed by  marked  metabolic  and  developmental  changes,  notable  being 
adiposity,  initritional  changes  in  the  skin  and  its  appendages,  sexual 

23  Arch.  Int.  Mod.,    100!)    (3).  4.3S. 

24  Praftitionor.  VMVA    (71).  472. 

2-.  Uculonmiillcr  iiiid  Stolt/.enbcrfr,  BiocliPin.  Zcit..  1010   (2S).  138. 

31  l''ull  l)il)]ioLn'a|)li\'  in  tlio  iii<)ii();,n"apli  bv  TTarv(>\'  Cushiiifr.  "Tlio  Piiiiii:u\  Podv 
and  ils  Disorders,"'  IMiiladclpliia,  1012 :'  also  .\sclni.T.  Ptliii;or"s  .Vrcli.'.  1012 
(]4(i),  1. 

32  Wells,  Jour.  Biol,  flicin..   1010    (7).  2.-)0. 

3-tT>(>\vis.  :\lillor  and  IMatllunvs.  Arch.  Tnt.  IMod..  1011  (7),  7S.-) :  llerriujr.  Quart. 
Jour.  Kxp.  Phvsiol.,  1014   (S).  24")  and  207. 

30  Sec  also   Pull.   .lolms   Hopkins'  IIosp.,    1013    (24).   40. 


Till-:  in  I'oi'in  s/s  .i\/>  ACh'oMjyi.iiA'  &15 

inaclivity  ami  undL'rdevL'lopment,  .subiKjnnal  body  temperature  ami 
increased  carbohj'drate  tolerance.^'^  These  manifestations  correspond 
to  those  observed  in  certain  Innnan  conditions  (Froehlicli's  syndrome) 
associated  witli  defects  in  tlie  hyi)opliysis.  Removal  of  the  posterior 
lobe  does  not  produce  any  characteristic  and  constant  effects,  altliongh 
marked  polyuria  and  erotism  have  resulted.  The  anterior  lobe  fed  to 
3^oun^  rats  has  a  stimulating  effect  on  growth,  and  especially  on  sexual 
development  and  activity,  while  posterior  lobe  feeding  has  a  retarding 
influence  (Goetsch^^).  Robertson  describes  a  modification  of  growtli 
in  mice  fed  anterior  lobe  substance,  which  he  attributes  to  a  specific 
substance,  tethelin,  containing  phosphorus  and  probably  an  iminazolyl 
group,  and  hence  related  to  the  active  constituent  of  the  posterior  lobe, 
although  it  has  no  pressor  effect. ^^ 

Puncture  of  the  hypophysis  produces  the  same  effect  as  puncture 
of  Bernard's  diabetic  center  in  the  fourth  ventricle,^^  and  stimulation 
of  the  glalid  has  a  similar  effect,  presumably  because  of  the  secretion 
of  a  glycogenolytic  agent.  A  diminution  of  posterior  lobe  secretion 
occurring  in  certain  conditions  of  hypopituitarism  leads  to  an  acquired 
high  tolerance  for  sugars,  with  the  resultant  accumulation  of  fat.  In 
hibernating  animals,  also,  the  adiposity  and  lowered  temperature  are 
associated  with  hypoplasia  of  the  anterior  lobe  of  the  hypophysis,  ac- 
cording to  the  same  author.  There  also  seems  to  be  some  relation  be- 
tween the  hypophj'sis  and  urinary  secretion,  for  extracts  of  the 
posterior  lobe  cause  marked  polyuria,  and  in  some  instances  of  "dia- 
betes insipidus,"  lesions  have  been  found  in  the  hypophysis.  Sim- 
monds  ^^  holds  that  the  pars  intermedia  is  responsible.  Like  the 
thyroid,  the  hypophysis  enlarges  during  pregnancy.^"  Feeding  of 
hypophysis  is  said  to  increase  both  gaseous  and  nitrogenous  metab- 
olism, and  in  a  case  of  hypopituitarism  the  urine  has  been  found  to 
contain  a  high  proportion  of  undetermined  nitrogen  and  of  neutral 
sulphur."'^  Varying  results  have  been  obtained  in  studies  on  the  basal 
metabolism  of  hypopituitarism.^^'' 

Acromegaly. — The  accumulating  evidence  seems  to  have  practically 
proved  that  acromegaly  depends  upon  a  hyperfunctionating  of  the 
anterior  lobe  tissue  of  the  hypophysis,  one  of  the  most  imjxn-tant  facts 
being  the  improvement  which  has  followed  removal  of  the  hyper- 
plastic tissues  in  several  cases  successfully  operated.  Although  there 
are  many  cases  of  tumor  of  the  hypophysis  without  acromegaly,  this 
is  of  no  significance  since  it  is  not  to  be  expected  that  all  tumors  will 

3T  Conoernino:  motaholism  after  hvpophvscctoniv  see  Benedict  and  Ilonians,  Jour. 
Med.  Pvcs.,  1012    (-2.")).  40<). 

33,Tohns  Hopkins  Hospital   Bulletin,   1016    (27),  20. 
35, Jour.  Biol.  ChenK.  1!»16    (24).  400. 
38Amer.  .Tour.  PliYsioI.,   1013    (.31),  xiii. 
39Mnneh.  med.  Woeli..   1013    (60),   127. 

40  See  Krdheini  and  Stumnie.  Ziegler's  Beitr.,   1000    (40).   1. 

41  Stetten  and  Rosenbloom,  Proc.  Soe.  exp.  Biol,  and  ^Med..  1013    (10),  100. 
41a  Means,  Jour.  Med.  Res.,  101.5    (32),  121. 


616  CHEMICAL    PATHOLOGY    OF    THE    DLCTLE^^    GLAyOfi 

carry  on  tlie  functions  of  the  tissue  in  which  they  arise.  Acromegaly 
without  hypoi)hyseal  changes  is  rare,  especially  if  we  consider  the 
liner  cytological  evidence  of  cellular  activity/-  So  far,  little  of  chem- 
ical interest  has  been  learned  concerning  this  disease.  The  metabo- 
lism studies  generally  indicate  a  retention  of  nitrogen,  phosphorus  and 
calcium,  because  of  the  overgrowth  of  bone  and  soft  tissues.^--'  Ac- 
cording to  some  observers  this  retention  is  decreased,  or  changed  to  an 
excess  elimination,  by  administration  of  hypophyseal  substance. ^^  The 
elimination  of  endogenous  uric  acid  is  said  to  be  greatlj^  increased 
in  acromegaly,  and  decreased  in  eases  with  hypofunction  of  the 
gland. ^*  A  considerable  excretion  of  creatine  was  'observed  by 
Ellis.-'-^ 

Glycosuria  and  actual  diabetes  is  frequently  present  in  acromegaly 
(40  per  cent,  of  the  cases  collected  by  Borchardt),*"  presumably  from 
interference  with  the  regulating  function  of  the  hypophysis,  but  this 
assumption  has  been  questioned  because  of  the  fact  that  lesions  in 
this  location  might  also  produce  glycosuria  by  affecting  the  "diabetic 
center."  However,  since  puncture  of  the  hypophysis  causes  glyco- 
suria, while  injection  of  posterior  lobe  extract  produces  glycosuria 
dependent  upon  hyperglycemia  (Gushing),  and  in  view  of  the  fact 
brought  out  by  Borchardt  that  in  cases  of  tumor  of  the  hy})ophysis 
without  acromegaly,  glycosuria  has  never  been  observed,  there  is 
much  probability  that  in  many  if  not  all  of  the  cases  of  glycosuria  with 
acromegaly,  it  is  the  liypophysis  itself  that  is  concerned,  and  that  both 
the  acromegaly  and  the  glycosuria  are  caused  by  hyperactivity  of  the 
gland. 

In  later  stages  of  acromegaly  there  may  develop  a  hypoactivity 
because  of  pressure  upon  the  posterior  lobe  or  infundibular  stalk, 
whereupon  the  sugar  disappears  and  is  replaced  by  an  increased  toler- 
ance for  sugar.^^ 

THYMUS-':   AND   OTHER  DUCTLESS  GLANDS 

From  the  clicrnical  stand|)()iiit  little  of  iiiterost  is  known  coneerning  this  orpan. 
Tt  is  frequently  used  as  a  souree  of  nucleic  acids,  in  which  it  is  rich,  but  there 
is  no  study  of  its  clieniical  clianges  tliat  is  of  interest  in  i>atholoufy.  l-Alirpation 
of  the  thymus  in  youiii;'  animals  is  followed  by  marked  defects  in  tlie  (le\('loi)inent 
of  the  skeleton,  and  changes  in  tlie  development  of  the  sex  organs.-''"  Some  au- 
thors state  that  thymus  extirpation  causes  a  loss  of  calcium,  and  tlial   a  calciinn 

42  See  Lewis,  'Bull.  Johns  Hopkins'  TTosp..  1005    (16),  InT. 

42;i  See  Hergeim,  Stewart  and  lii'wk.  Jour.  Exp.  ^led.,   ini4    (20).  21S. 

4^  See  Kidiinraiit,  Dissert.,  Zuricl',  (Jebr.  l.ceman,  1012:  Mediureccanu  and 
Kristeller,  .Tour.  Biol,  ("hem.,   101 1    ( !) ) ,   100. 

■"Falta  and  Nowaczynski,  Berl.  klin.  Woch..   1012    (40),   ITSl. 

■'•"'Jour.  Amer.  Med.  Assoc.   1011    (5(>),    1S70. 

••«  Zeit.  klin.  Med.,  lOOS    (fifi),  X\2. 

••7  Full  discussion  in  .Johns  Hopkins  TIosp.  Bull.,  1011    (22).  M\'->:   101:?   (24),  40. 

2"  In  addition  to  Biedl's  "Innere  Sekretion."  see  Wiesel,  Krgebnisse  Plnsiol., 
1011  (XV  (,)  ),  41(>:  Klose  and  Vogt,  Beitr.  z.  klin.  Chir.,  lOlO  ((10),  1;  Matti, 
.Mitt.  (Jrenz.  .Med.  u.  Chir..   1012    (24),  H.  4-5. 

2"ia  Not  corroborated  iiv  Taiipcnlieimer  (.Tour.  Kx]).  Med..  1014  (10).  :nO;  (20), 
477)   or  by  Nordmann,    (Ardi.  klin.  Ciiir..   1014    (KH!),   172). 


TUYMVH  AM)  o'lHEIi  DUCTLESS  GLAyDS  617 

rotentioii  results  fidiii  feeding'  tliviiius,  Iml  the  ri'sulls  (|iintc(l  ari'  not  at  all  in  liar- 
mony.  It  is  certain,  liowovcr,  tiiat  (h'lici(>n<-y  in  tlie  tliynuis  causes  a  defect  in 
ossification.  Also  llierc  occurs  a  period  of  adiposity,  followed  hy  cacliexia  witli 
liyperplasia  of  tiic  lynijihatic  tissues,  thyroid,  pancreas,  ovaries  and  testicles 
(  Klose  and  \'o','t ) .  These  authors  attrilnite  the  defects  in  ossification  to  aii 
acidosis,  which  liypothesis  is,  liowever.  far  from  established.  ( ludernatscli  27 
found  that  the  fecdin<r  of  tliymus  to  tadpoles  causes  a  great  increase  in  the  rate 
of  growth,  and  decreases  or  suppresses  the  develoj)mental  changes,  liaving  exactly 
the  opposite  efTect  from  tliyroid  feeding,  and  Abderhalden  2Ta  liag  found  that  this 
property  persists  after  digesti(m  of  tlie  thymus  tissue.  As  yet  no  substance  has 
been  isolated  which  can  be  considered  as  a  sjjccific  internal  secretion  of  the 
thymus,  altliougli  the  frc<|uent  conciu-rence  of  abnornuil  conditions  in  the  thyroid 
and  thymus,  in  the  adi^cnals  and  thymus,  in  the  hypophysis  and  thymus,  to- 
gether with  tlie  frei|uency  of  ])olyglaii(lular  conditions,  leaves  no  (|ucstion  that  the 
thymus  is  to  be  considered  with  tlic  other  members  of  this  system,  however  differ- 
ent its  histological  structure  may  be.^Tb  Thymus  administration  by  mouth  does  not 
counteract  the  loss  of  the  thymus.  The  enlargement  of  tlie  thymus  that  occurs  in 
most  cases  of  exophthalmic  goiter  is  accompanied  at  times  by  symptoms  that 
suggest  an  intoxication   from  this  source.27c 

The  chemistry  of  tlio  Pinfal  Glaxd  can  be  dismissed  practically  without  con- 
sideration, since  no  positive  facts  have  lieen  brouffht  to  light.  Extracts  from  the 
organ  show  no  distinct  physiological  eflfects.^s  Tumors  of  the  pineal  gland  have 
been  found  associated  witli  adiposity  and  witli  precocious  sexual  devel<i])nient,  but 
whether  from  the  action  of  the  gland  itself  or  from  the  pressure  on  the  brain, 
cannot  be  said. 29  Extirpation  of  the  pineal  seems  to  have  no  noticable  effects  of 
any  sort  (Dandy  2na)  although  ^IcCord  29b  reports  increased  growth  and  early 
sexual  maturity  in  animals  fed  pineal  substance. 29c 

The  Carotid  Glaxd  is  more  directly  related  to  the  adrenal  medulla,  in  that  it 
contains  chromaffin  tissue.  It  should,  presumably,  contain  a  pressor  principle,  as 
Moulon  found,  l>ut  Gomez  30  obtained  only  lowering  of  blood  pressure  fi-om  extracts 
of  this  gland,  and  bilateral  removal   causes  no  characteristic  effects.soa 

2T  Arch.  Entwicklgs.,  ini2    (35),  457:   Amer.  Jour.  Anat.,  1914    (15),  431. 

27a  Arch.  ges.  Physiol.,  1915    (162),  99. 

27b  Literature  given  by  Basch,  Zeit.  exp.  Path.,   1913    (12),   180. 

270  See  review  by  Halsted,  Bull.  Johns  Hopkins  Hosp.,  1914    (25),  223. 

28  Jordan  and  Eyster,  Amer.  Jour.  Physiol.,  1911  (29),  115:  Dixon  and  Halli- 
burton, Quart.  Jour.  Exper.  Phvsiol.,  1909  (2),  283.  Dana  and  Berkelev.  Med 
Record,   1913    (S3),  Xo.   19. 

29  See  Pappenheimer,  Virchow's  Arch.,  1910    (200),   122. 
29a  Jour.  Exp.  :Med.,   1915    (22),  237. 

29b  Jour.  Amer.  Med.  Assoc,  1914    (63),  232  and  517. 

29c  Concerning  the  composition  of  the  pineal  gland  see  Fenger,  Jour.  Amer.  ^led 
Assoc.  1916    (67),  1836. 

30  Amer.  Jour.  IVIed.  Sci.,  Julv,  1908. 

30a  Massaglia,   Frankf.   Zeit.   Path.,    1916    (18),  333. 


CHAPTER    XXI 

URIC-ACID  METABOLISM  AND  GOUT  ' 

These  subjects  have  been  the  o])jeet  of  such  a  prodigious  amount 
of  research  that  it  is  far  beyond  the  seope  of  tliis  work  to  review  the 
histor}'  and  the  details  of  the  investigations.  Such  a  review  is  also 
particularly  unnecessary,  since  it  can  be  found  in  the  works  on  phys- 
iological chemistry  and  various  treatises  on  metabolism.  Conse- 
quently the  attempt  will  be  made  in  this  chapter  merely  to  give,  as 
brietiy  as  possible,  the  views  now  most  generally  accepted  concerning 
the  nature  and  metabolism  of  uric  acid,  and  its  relation  to  patho- 
logical processes.  For  the  historical  discussion,  indicating  by  what 
devious  steps  we  have  reached  our  present  understanding  concerning 
this  long-disputed  subject,  the  reader  is  referred  to  the  articles  men- 
tioned below,  upon  which  I  have  drawn  freely.  A  particularly  clear 
summary  of  the  subject  is  given  by  Walter  Jones  in  his  monograph  on 
nucleic  acids.^ 

THE  CHEMISTRY  OF  TTRIC  ACID 

It  is  the  very  great  service  of  Emil  Fischer  to  have  shown  us  the 
structure  of  the  uric-acid  molecule,  the  empirical  formula  of  which, 
C3H4N4O3,  had  long  been  known.  He  demonstrated  that  it  is  a  mem- 
ber of  a  group  of  substances,  which  are  all  characterized  by  being 
built  up  about  a  certain  nucleus,  C-N^.  As  the  simplest  member  of 
the  group  is  a  synthetically  formed  body,  purine,  the  nucleus  is  called 
the  "purine  nucleus."  The  structural  relations  of  the  better-known 
"purine  todies"  to  this  purine  nucleus  and  to  each  other  is  clearly 
shown  by  their  structural  formula?,  as  given  below : 

The  atoms  in  the  "purine  nucleus"  are  arranged  as  follows: 

N(i)— C(6) 

I  I 

C(2)— C(5)— N(t) 
N(3)-C(4)-N(9) 

To  each  atom  has  been  given  a  inimber,  as  shown,  for  the  purpose  of 
facilitating  reference  to  the  location  of  various  atoms  and  groups 
that  are  attached  to  this  nucleus.  The  structure  of  i)uriiu'  itself  is 
as  shown  on  the  following  page :  - 

1  CompU'te  rpviows  arp  irivcn  l)y  1'.  TI.  .McCniddcii,  "rric  At'id,"  Xcw  ^Orlc,  lOOn; 
Wionor,  Krpol)nissf  dor  Plivsiol.,"  1002  (1),  fj;");") ;  ibid.,  l!t().S  (2),  :?77;  Burian  and 
Sdmr,  Pfliifrcr's  Arcli.,  ino'o  (SO),  241;  1001  ( S7 ) .  2:5!);  Sc'liiltonludni.  Ilaiidh.  d. 
Tliocliciii..  1010,  IV  (,),  4S0;  Bru'-srh  and  Scliittcnholm,  "Die  NukleinstolTwoHisel 
und  seine  St.r)run<,'('n,"  .Jena,  1010;  Waller  -lones,  "Niudeie  Acids,"  Monograplis  on 
JJioclieniistrv,  1014.  An  excellent  sinnniai\-  of  i-eccnt  woik  is  i;iven  bv  Benedict, 
Jour.  l.ab.  Clin.  Med.,  lOlG   (2),  1. 

-In  these  formuhe  the  symbols  of  llie  atoms  foiining  tiie  purine  nucleus  are 
in  heavy  type. 

618 


THE  CHEMISTRY  OF  URIC  ACID  619 


N=CH 

I       I 
HC    C-NH 

II    II    X 


CH 


N-C-N 

Purine 


The  derivatives  of  purine  are  described  by  stating  to  which  atom 
of  tlie  purine  nucleus  the  combining  groups  are  attached.  Thus, 
adenine  is  referred  to  as  6-aniino-purine,  and  therefore  has  the  follow- 
ing formula : 

N=C-NH2 

I  I 

HC    C— NH 

II  II       r^" 
N-C-N 

Adenine  (6-amino-purine) 

Other  important  members  of  this  group  of  "purine  bodies,*'  (also 
called  xunthine  bodies,  alloxuric  bodies,  and  nuclein  bodies)  are  built 
up  about  the  purine  nucleus  as  shown  below : 

HN-C=0  HN-C=0 

II  I      I 

H2N.C  C-NH  0=C  C-NH 

II  II  >^»  i  II  >^"- 

N-C-N  HN-C-N 

Guanine  Xanthine 

(2-aniinoUoxypurine)  (2,  6-dioxypurine) 

HN-C=0  HN-C=0 

II  II 

HC    C-NH  0=C  C-NH 

II  II  ^^"  i  II  /*^=^- 

N-C-N  HN-C-i^H 

Hvpoxanthine  Uric  acid 

(6-oxypurine)  (2-6-8trioxypurine) 

H3C-N-C=0  H>J=C=0 

I       I  .CHa  I       I  CH3 

o=c  c-n/  o=c  C-N/ 

!    II    ^cH  !    II    >^" 

H3C-N-C-N  H3C-N-C-N 

Caffeine  Theobromine 

(1-3-7  trimethyl-2-6  dioxypurine)  (3-7-dimethyl,  2-ti  di.ixypurine) 

As  shown  by  their  structural  formula?,  the  purimidinrs  present  in 
the  nucleic  acids  are  also  closely  related  to  the  ])urines,  viz: 

N  — CH  X  — C  — NH=       HN  — C  =  0      HX  — Cc=.0 

HO   CH       0  =  C   CH         0  =  C   CH      0<=.C   C  — CH» 

N  =  CH         HN^  — CH  TIN-  — CH        HN  — CH 

Pvrimidine                               Cvtosine  Uracil                                  Thymine 

(2-oxy.  6-animo-  (2-6-dioxy-  (5-methyl.  2-6-dioxy- 

pyrimidine)  pyrimidine)                        pyrimidine). 


620  CRIC.WID    METABOLISM    AXD    GOUT 

Properties  of  Uric  Acid. — Uric  acid,  when  pure,  is  white,  and 
crystallizes  in  rhombic  tablets.  Its  solubility  is  very  slifi'ht ;  at  room 
temperature  (18°)  it  dissolves  but  about  one  part  to  40,000  of  water, 
so  that  a  saturated  solution  contains  but  0.0253  gram  to  the  liter. 
It  is  much  more  soluble  in  blood-serum,  dissolving  in  1000  parts,^ 
])robably  held  in  some  complex  combination.  His  and  Paul  have 
shown  that  in  a  saturated  solution  oidy  9.5  per  cent,  of  the  molecules 
are  dissociated,  tlie  dissociation  occurring  in  two  steps;  the  Urst  and 
chief  dissociation  is  into  H  and  C.,H..5N^0.j,  which  then  undergoes  fur- 
ther dissociation  into  H  and  Cr.HoN^O.,,  the  latter  dissociation  being 
very  slight.  If  any  other  acid  is  present  in  the  solution,  its  dissocia- 
tion and  liberation  of  free  hydrogen  ions  interferes  with  the  dissocia- 
tion of  the  uric  acid,  and  as  the  undissociated  uric  acid  is  extremely  in- 
soluble, the  amount  dissolved  in  an  acid  solution  is  much  less  than  in 
a  neutral  solution.^''  Gudzent  *  found  that  saturated  solutions  of 
urates  gradually  precipitate  out  the  salts  because  of  a  transformation 
of  part  of  the  uric  acid  into  what  he  believes  to  be  a  I-actim  form. 
(The  lactim  form  is  shown  in  the  following  formula,  as  compared  with 
the  isomeric  lactam  form  shown  above,  in  which  uric  acid  is  supposed 
to  exist  ordinarily.) 

N  =  C  —  OH 


HO  — ( 

fi- 
ll 

■NH 

\ 

r- 
// 

—  OH 

] 

<!■ 

—  c  — 

-NH 

(Lacti 

m 

form  of 

uric  a 

cid) 

With  alkalies  uric  acid  yields  two  series  of  salts,  corresponding 
to  these  two  steps  in  dissociation :  one,  in  which  one  atom  of  the  base 
enters,  is  called  the  hiurate  or  monohasic  urate ;  the  other  is  the  so- 
called  ''neutral"  or  hihasic  urate.'''  Of  the  two,  the  latter  is  much 
the  more  soluble.  The  monosodium  urate  forms  colloidal  solutions 
in  water,  from  which  the  crystalline  salt  gradually  falls  out.  The 
quadriurate,  of  which  much  was  said  in  the  earlier  literature,  prob- 
ably does  not  exist  (Kohler).*' 

In  the  urine  the  uric  acid  and  the  urates  are  kept  in  solution  by 
the  pliosphates,  the  disodiuui  phosphate  preventing  the  decomposition 
of  tlie  urates  into  uric  acid  by  the  acid  salts  of  the  urine.  Possibly 
other  constituents  of  the  urine,  especially  the  pigments  and  NaCl,  also 
aid  in  its  solution.  Urine  may  form  quite  stabl(»  supersaturated  solu- 
tions and  Kohler  states  that  the  urine  is  a  ti'uly  suix'rsaturated  solu- 

s  Taylor,  Jour.  Biol.  Cheni.,  IDOG    (1),   177. 

3a  Concorniiitr  tlio  sohibilitv  of  uric  acid  in  urine  sec  I[a>kiiis,  .lour,  liiol.  Clieiii., 
I'tlO    (26),  205. 

«Z('it.  pliysiol.  Chcin.,   I'KI!)    ((iO),  .38. 

•'■' .Xs  <'i.  matter  of  fact,  l)olli  salts  ^ive  a  sli<ililly  alkaline  reaction  wlien  dis- 
Holved   in  water    ('i'a\lor). 

'■Zeit.  physiol.  Cliom.,   1!)11    (72),  10!);    1!)13    (78),  205. 


i'(nru.\'i/o\   ()!■'  I  h'lc  .\<ii)  621 

tion  of  sodiiuii  urate.  How  the  uric  acid  is  kept  in  solutiiui  in  tile 
blood  is  not  exactly  understood,  but  Gudzent  believes  that  uric  acid 
can  exist  in  the  blood  only  as  the  monosodiuni  urate,  and  in  the  less 
soluble  but  more  stable  lactim  form,  which  is  soluble  only  to  the  extent 
of  8.3  mp;.  per  100  ec.  serum  (the  lactam  foi-m  beinj^  soluble  up  to  18 
mg.).  However,  amounts  over  20  m<z:.  i)er  100  c.c.  have  been  detected 
in  the  blood  of  nephritics;  here  solution  may  have  been  aided  by  the 
other  retained  metabolites.  Bechhold  "^  and  others  have  maintained 
that  urates  may  be  present  in  the  blood  in  a  colloidal  state  which -can- 
not pass  out  throu^li  the  kidneys. 

FORMATION   OF  URIC  ACID  " 

The  origin  of  uric  acid  is  chiefly,  although  not  exclusively,  from 
the  nucleoproteins,  and  it  is  customarA'  to  refer  to  uric  acid  formed 
from, the  nucleoproteins  of  the  foods  as  "exogenous'^  uric  acid,  in  con- 
trast to  the  " endogenous"  wr'xQ,  acid  that  is  formed  from  the  nucleo- 
proteins of  the  body  cells  during  their  catabolism.  This  may  be  read- 
ily explained  hy  a  brief  consideration  of  the  composition  of  the  nucleo- 
proteins. The  nucleoproteins  may  be  looked  upon  as  salts  formed 
through  combination  of  proteins  with  nucleic  acid.  Nucleic  acid  in 
turn  is  a  compound  of  phosphoric  acid  with  purine  bases,  pyrimidine 
bases,  and  carbohydrate  radicals,  constituting  a  complex  sort  of  glu- 
coside. 

A  long  series  of  careful  analytical  studies  has  at  last  shown  us  that 
nucleic  acids,  are,  whatever  the  source,  quite  similar  in  composition, 
consisting  always  of  a  complex  containing  phosphoric  acid,  the  two 
amino  purines  (adenine  and  guanine),  two  pyrimidines  (either  cyto- 
sine  and  uracil  or  cytosine  and  thymine)  ;  and  a  carbohydrate,  which 
may  be  either  a  pentose  or  a  hexose.  Apparently  there  are  two  sorts 
of  nucleic  acids,  one  from  plants,  which  contains  always  uracil  and 
pentose,  and  one  from  animal  tissues,  containing  instead  thymine  and 
a  hexose.  So  constant  are  the  findings  in  regard  to  these  compounds 
that  it  has  seemed  feasible  to  consider  their  manner  of  union  in  the 
intact  nucleic  acid  molecule,  and  Levene  and  Jacobs  have  ]n-oposed  as 
the  structure  of  thymus  nucleic  acid  the  following  arrangement : 

H    POs  —  CeH.oOi  —  CsHsNeO 

!  (guanine  sroup) 

? 

U=P04  —  CeHsOa  —  CsHbN^O: 

1  (thymine  group) 

? 

H2PO4  —  CeFs02  —  C4H4N3O 

I  (cvtosine  group) 

o 

H   PO3  —  CeH-oO*  —  C5H4N5 

(adenine  group) 

saBiocliem.  Zeit.,  1914   (64),  471. 

"See  review  in  International  Clinics,  1910,  XX    (J,  76. 


622  URIC-ACID   Mf-rri HOLISM  axd  gout 

It  will  be  seen  that  this  proposed  formula  postulates  in  the  nucleic 
acid  molecule,  one  radical  of  each  of  the  two  purines  and  pyrimidines, 
eacli  of  these  ho'mg  iniited  by  a  carbohydrate  radical  to  a  i)hosphoric 
acid  radical.  Reco<:-nizing  that  tliis  must  be  looked  upon  as  a  provi- 
sional fonnula,"''  it  will  serve  as  a  base  of  departure  from  which  to 
consider  the  metabolism  of  nucleic  acid. 

The  groupin<r  of  hexose  -\-  purine  or  hexose  -|-  pyrimidine  is  re- 
ferred to  as  a  "luicleoside,"  analogous  in  terminology  to  "glueoside." 
The  same  groupings  plus  the  phosphoric  acid  radical  constitute  the 
"nucleotids, "  nucleic  acid  thus  being  made  up  of  four  nucleotids. 
Emil  Fischer  has  reported  ~^  the  synthetic  production  of  a  nucleotid, 
composed  of  pliosj^horic  acid  united  to  a  glucoside  of  theophyllin,  this 
really  constituting  the  long-sought  synthesis  of  a  nucleic  acid,  even 
though  the  artificial  product  is  not  the  same  as  any  known  to  occur  in 
nature.  With  these  facts  before  us  we  may  consider  the  manner  in 
which  nucleic  acids  are  disintegrated  in  the  animal  body. 

So  large  a  molecule  can  conceivably  be  disintegrated  in  many  differ- 
ent ways ;  that  is.  the  lines  of  cleavage  might  pass  through  several 
different  points  and  in  many  different  orders,  but  there  is  evidence 
available  which  causes  us  to  believe  that  the  process  is  quite  constant 
in  animal  metabolism.  Jones  considers  it  probable  that  the  first  step 
is  a  decomposition  of  the  tetranucleotid  into  dinucleotids,  and  that 
these  are  in  turn  split  into  mononucleotids.  Little  is  known  about  the 
subsequent  career  of  the  two  pyrimidine  nucleotids,  but  we  have  an 
abundance  of  information  concerning  the  nucleotids  containing  the 
purines,  and  it  is  in  these  our  present  interest  lies.  Each  nucleotid 
has  two  points  at  which  it  might  be  split,  and  we  have  reason  to  believej 
that  there  exist  in  animal  tissues  enzymes  which  may  specifically  at- 
tack each  bond.  One  enzyme  separates  the  pliosphoric  acid  radical 
from  the  nucleoside,  thus :  | 

HjPO,  -  C.HA  -  C,H,N50  +  H.O >- H^PO,  +  C,,H ,0,  -  C,H,N,0 

guanylic  acid  phospho-nuclease  guanosine 

and  this  enzyme  is  therefore  designated  as  jjhospho-nuclcasc. 

Another  enzyme,  purine  nuclease,  splits  off,  instead,  the  purine  radi- 
cal, thus: 

HjPO,  -  C,H,03  -  r,H,N,0  +  11,0 y  ILPO,  -  C,HA.  +  C.HiN^O 

guanylic  acid  i)uriiie-nuclease  guanine 

Following  either  of  these  cleavages,  the  enzymes  which  deaminize 
l)ui-ijies  ])egin  to  act,  and  we  have  formed  as  a  result  either  the  free 
oxypurines  or  the  oxy purines  still  boimd  in  the  glucoside-like  combi- 
nation with  sugar.     If  the  purines  are  free  the  reaction  will  be: 

■:>  Jdiu's  and  ITcad  (Jour,  l^iol.  Clicni.,  1!)17  (2!>),  11])  liavo  advanced  evidence 
to  indicate*  that  tlie  liid<af,'e  between  tlie  neiiclent  ids  is  between  the  earhojiydrate 
radicals  rather  than  between  the  ])h(»s|)]u)ric  acid  jri^onps. 

7b  Sitzungsbcr.  k.  Akad.  Wissensch.,  J^erlin,  1*114   (IJ.'?),  DOf). 


i'()h'M.['H(>.\  OF  I  inc  Ar/n  023 

C\.II,N,0  +  ILO >-C',n,X,(),,  +  XII3 

guanine  ;;uaua!!K         xantliiiiu 

or,  hi  case  the  guanine  glueoside  is  present: 

C5HA-C,H4N„0  +  H,0 >- CJIA  -  C,H3N  A  +   ^ih 

suanosinc  guanosinc-deaniinasc  xanthosine 

lu  the  latter  case  a  hydrolytic  euzynie,  xaiithosine-hydrolase,  then 
splits  off  the  xanthine,  so  that  by  either  route  the  end  result  is  the 
same.  By  a  similar  series  of  changes  the  adenine  radical  is  converted 
into  hypoxanthine,  either  directly  by  adenase: 

C5H3N,  +  ILO >-Cjr,N,0  +  XH3 

adenine  aden;:se  h\  poxanthine 

or   by   adenosine-deaminase   the   hypoxanthine-glucoside    (inosine)    is 
formed,  and  later  the  hypoxanthine  is  split  off. 

We  now  have  hypoxanthine  and  xanthine,  which,  in  the  presence  of 
oxygen,  are  oxidized  to  form  uric  acid,  thus : 

QH^N.O  +  O ^ >^C,H,N,Oo 

hypoxanthine     hypoxanthine-oxidase       xanthine 

CH^NA  +  O y  C-,H,N,03 

xanthine  xanthine-oxidase  uric  acid 

Further  oxidation  of  the  uric  acid  causes  its  conversion  into  the 
much  more  soluble  allantoin,  thus : 

C5H,N,03  +  0  +  H,0— >-C,HeN,03  +  CO^ 

uric  acid  uricase  allantoin 

It  is  thus  evident  that  the  steps  of  the  disintegration  of  nucleic  acid 
are  numerous,  but  that  each  separate  process  is  a  simple  one ;  and  also, 
that  it  has  been  possible  to  follow  out  and  distinguish  the  several  steps 
and  to  establish  the  fact  that  each  step  depends  on  a  distinct  and 
specific  enzyme.  Not  every  tissue  possesses  all  the  enzymes  of  purine 
destiniction,  and  in  different  species  of  animals  the  distribution  of  the 
enzymes  is  different.  For  example,  the  enzyme  xanthine-oxidase, 
which  oxidizes  xanthine  into  uric  acid,  is  found  in  man  only  in  the 
liver,  and  also  in  other  animals  it  is  of  limited  distribution,  being 
found  usually  only  in  the  liver  or  in  the  liver  and  kidney,  but  in  the 
dog  it  seems  to  be  present  in  several  tissues.  The  deaminizing  en- 
zymes, adenase  and  guanase,  are  much  more  widely  distributed,  but  by 
no  means  universally.  Adenase,  for  example,  is  not  present  in  the 
tissues  of  the  rat,  and  not  in  the  tissues  of  adult  human  beings.^ 
Guanase  is  absent  from  the  spleen  and  liver  of  the  pig  and  from 
human  spleen,  although  present  in  most  other  tissues.  Uricase,  the 
enzyme  which  destroys  uric  acid,  also  has  peculiarities  of  distribution, 
being  seldom  found  in  any  other  tissue  than  the  liver  or  kidney,  and 
being  absent  entireh-  from  the  tissues  of  man,  and  from  the  birds  and 

8  There  liave  l)cen  some  rejiorts  indicatiiisj  tlie  presence  of  adenase  in  fetal  liuman 
tissues   (Long,  Jour.  Biol.  Chem.,  101.3   (la),  440). 


624 


URIC-ACID    METABOIJ^M    A\D    GOUT 


reptiles  so  far  examined.     The  significance  of  this  distribution  of  iiri- 
case  will  be  discussed  at  greater  length  a  little  later. 

The  following  graphic  expression  of  the  series  of  steps  leading  to 
the  formation  of  uric  acid  has  been  presented  by  Amberg  and  Jones:'' 


0--P'-Q  CsHjOj      Cs  HjN,  (NHj) 


^u.n.n 


Uric  add 

/OH 

C,HN-OH 


oxidcie     Xanthine    ^ 


adenosine 


[anrhlne 


ypm, 
C.HjN^O  H 


Another  possible  source  of  uric  acid  is  through  synthesis.  In  birds, 
which  eliminate  most  of  their  nitrogen  in  the  form  of  uric  acid,  syn- 
thesis of  uric  acid  undoubtedly  occurs.  It  must  also  be  considered 
that  young  mammals  can  synthesize  the  purines  necessary  for  their 
growth  from  foods  which  contain  no  purines.^*'  It  would  seem  pos- 
sible, therefore,  for  synthesis  of  uric  acid  to  occur  in  adult  mammals, 
but  as  yet  satisfactory  experimental  evidence  is  lacking  that  such 
synthesis  does  occur,  although  an  apparently  reversed  reaction, 
whereby  uric  acid  destroyed  by  liver  tissue  can  be  resj^nthesized  by 
the  same  tissue  acting  upon  it  in  the  absence  of  oxygen,  has  been  de- 
scribed by  Ascoli  and  Izar.^^  Their  work  has  not  been  repeated  suc- 
cessfully by  others.  I  have  failed  in  several  attempts  to  secure  re- 
synthesis  of  uric  acid  by  dog  livers,  and  Spiers/^''  who  made  a  more 
extensive  investigation,  was  unable  to  corroborate  their  findings. 

It  should  also  be  mentioned  that  not  all  of  the  purine  bases  of  the 
body  is  bound  in  the  form  of  nucleic  acid.  A  considerable  amount  is 
present  in  a  free  condition,  or  at  least  not  bound  in  luicleic  acid,  espe- 
cially in  muscle  tissue.  Uric  acid  can  be  fonned  as  well  from 
the  free  purine  bases  as  from  purine  bases  liberated  from  nucleic  acid 
— indeed,  evidence  has  been  brought  forward  indicating  that  a  large 
proportion  of  the  uric  acid  arising  during  metabolism  (endogenous) 
comes  from  the  free  hypoxanthine  of  the  muscles. 

As  to  the  place  where  uric  acid  is  formed,  it  seems  probable  that  in 
different  animals  different  organs  arc   chiefly   coiu'erned.   for   it   has 

oZeit.  plivsiol.  Chvm.,  1011    (7.S),  407. 
lOMK'olluin,  .Amcr.  .lour.  Plivsiol..  100!)   (2r>).  120. 
11  Sf'c  Zeit.  physiol.  Clicin..  1010   (05),  78. 
iiaBiochem.  .lour.,   l!)l.'i    (0),  -.VM. 


BEST ix' I  cm )\  (>r  uuic  acid  625 

been  found  tliat  tlie  (listrihution  of  the  en/ymes  mentioned  above  varies 
jjreatly  in  tlie  various  organs  and  tissues  of  different  species/'-  In 
most  animals  tiie  xanthine  oxidase,  wliieli  forms  uric  acid  from  xan- 
thine, is  localized  chiefly  or  solely  in  the  liver,  and  this  is  the  case  in 
man ;  therefore  it  is  presumable  that  uric  acid  is  formed  chiefly  in  the 
liver  from  jnirines  hy  the  steps  described  above.  That  there  may  be 
otlier  metliods  of  foniiiiiu'  uric  acid  is  possible. 

DESTRUCTION  OF  URIC  ACID  i  '• 

With  most  mammals  but  little  of  the  total  amount  of  purine  bases 
taken  as  food  or  set  free  in  the  tissues,  appears  in  the  urine  as  uric 
acid,  most  of  it  being  converted  into  allantoin,  which  seems  to  be  ex- 
ereted  witli  little  or  no  loss.  Thus,  when  dogs,  pigs  or  rabbits  are  fed 
nucleic  acid,  about  93  to  95  per  cent,  can  be  recovered  as  allantoin, 
3  to  6  per  cent,  as  uric  acid,  and  1  to  2  per  cent,  as  purine  bases 
(Schittenhelm).  It  would  seem  that  practically  all  the  purines  can 
be  found  in  these  three  forms  combined,  the  proportions  varying  in 
different  species.  In  man  alone,  except  for  the  chimpanzee  ^*  and 
orang-utan,  does  a  considerable  proportion  escape  as  uric  acid,  a  fact 
in  complete  harmony  with  repeated  observation  that  the  tissues  of 
man  have  no  power  whatever  to  destroy  uric  acid  in  vitro;  the  earlier 
reports  of  positive  uricolysis  undoubtedly  being  erroneous.  Even 
the  monkey  has  active  uricolytic  enzymes  in  its  liver,  and  therefore 
excretes  its  purines  chiefly  as  allantoin.  With  mammals  as  a  whole, 
therefore,  uric  acid  is  destroyed  to  the  extent  of  being  converted  into 
allantoin,^*''  the  close  relationship  of  which  to  uric  acid  is  shown  by 
the  structural  formula: 

NH  —  PH  —  NH 

o  =  i^l— i  =  0 

I        I       I 

NH  —  CO  —  NH= 

(allantoin) 

With  most  mammals  the  oxidation  of  uric  acid  takes  place  chiefly  in 
the  liver,  but  in  some  of  the  herbivora  the  kidneys  are  more  active,  as 
far  as  experiments  in  vitro  can  show. 

Whether  man  can  destroy  uric  acid  at  all  has  been  a  matter  of  dis- 
pute. It  has  been  shown  by  Wiechowski  and  others  that  uric  acid 
injected  subcutaneouslj^  is  excreted  almost  quantitatively  and  un- 
changed in  the  urine.  To  be  sure,  human  urine  does  contain  a  very 
little  allantoin.  7  to  14  mg.  per  day,  but  this  amount  is  too  small  to  be 
of  much  significance,  for  it  is  possibly  all  derived  from  the  food,  as 

12  A  compilation  of  this  distribution  is  given  hv  Wells,  Jour.  PJiol.  Clieni.,  1010 
(7),  171. 

13  See  discussion  by  Wells.  Jour.  Lab.  Clin.  "Med.,  inio   (1).  104. 

1*  Wiechowski,  Pras-er  nied.  Woch..  1012  (37),  27o :  Wells  and  Caldwell.  .Tour. 
Biol.  Chem.,  1014    (18),  157. 

14a  See  Hunter  and  Civens,  .Jour.  Biol.  Chem.,  1914   (IS),  403. 
40 


626  VBICACID    METAHOLISM    A\D    aOT'T 

1lie  liuman  organism  cannot  desti-ny  allaiitoiii.^^''  On  tlu'  other  hand, 
it  lias  been  found  repeatedly  that  nneleie  aeid  or  purines  given  by 
mouth  are  by  no  moans  (|uantitatively  excreted  in  the  urine,  even 
when  uric  acid,  allantoin  and  purine  bases  are  added  together.  Ap- 
parently a  considerable  proportion  of  the  purine  nitrogen  fed,  about 
half  in  most  experiments,  is  excreted  as  urea.^*'=  As  allantoin  seems 
not  to  be  at  all  disintegrated  in  the  human  body  it  would  seem  prob- 
able that  if  purines  are  destroyed,  as  these  experiments  indicate,  they 
pass  through  some  other  route  than  allantoin,  and  possibly  that  part 
of  the  purines  which  is  destroyed  does  not  pass  through  the  stage  of 
uric  acid.  Experiments  show  that  outside  the  body  uric  acid  can  be 
destroyed  by  other  routes  than  through  allantoin;  thus,  it  can  be  dis- 
integrated into  glycocoll,  ammonia  and  CO2 ;  or  by  another  method  of 
destruction  it  yields  first  alloxan  (C4H2N2O4),  then  parabanic  acid 
(CjHoNjO.t)  which  in  turn  yields  oxalic  acid  and  urea.  There  is  no 
evidence,  however,  that  any  of  these  alternative  routes  is  ever  fol- 
lowed in  the  aiiimal  body.  It  is  possible  that  the  failure  to  find  all 
the  purines  of  the  food  as  uric  acid  in  the  urine  depends  on  their  par- 
tial destruction  in  the  intestine  by  bacteria.""^  It  is  highly  probable, 
in  view  of  all  available  evidence,  that  in  man  most  of  the  purine  ab- 
sorbed from  the  food,  and  practically  all  the  purine  from  cell  metabo-  ■ 
lism,  is  converted  into  uric  acid  and  excreted  as  such. 

THE   OCCTJRRENCE    OF   URIC   ACID   IN   THE   BLOOD,   TISSUES.    AND    URINE 

As  can  be  seen  from  the  foregoing  discussion,  the  amount  of  uric 
acid  that  appears  in  the  urine  depends  upon  a  number  of  factors, 
which  may  be  enumerated  as  follows:  (1)  The  amount  of  purine 
bodies  taken  in  the  food,  upon  which,  chiefly,  depends  the  amount  of 
exogenous  uric  acid.  (2)  The  amount  of  destruction  of  tissue  nucleo- 
proteins.  (3)  The  amount  of  purine  bases  formed  in  the  muscle 
tissue.  (4)  The  amount  of  conversion  of  purine  bases  into  the  uric 
acid.  (5)  The  amount  of  destruction  of  uric  acid,  if  any,  occurring 
in  the  body.  (6)  Possibly  upon  the  capacity  of  the  tissues  to  syn- 
thesize uric  acid ;  and  in  case  such  power  to  synthesize  uric  acid  exists, 
uDon  the  presence  of  the  precursors  of  uric  acid  in  the  body.  (7) 
The  retention  of  uric  acid  in  the  blood  and  tissues.  fS'i  The  power 
of  the  kidneys  to  excrete  uric  acid. 

If  we  also  take  into  account  the  fact  that  the  solubility  of  uric  acid 
in  the  urine  depends  chiefly  upon  the  amount  of  neutral  phosphates 
present  in  the  urine,  and  also  upon  the  temperature,  reaction,  and 

14b  See  Ac-krovd.  T.iodiciti.  .Tour.,  1011    (.T),  217,  400,  442. 

Hf"  Soe  Taylor.  Jour,  r.i.il.  Clioni..  101.3  (14),  410;  Hivons  and  TTuiitor.  ihld..  lOlf) 
(2.3),  200.  A1)ont  nno-toTilli  as  miieli  uri("  acid  is  ovcrclcd  in  tlic  swoat  as  in  tlio 
urine,  sweat  containinfr  0.1  ni<j;.  ptT  c.c.  Adlor,  Dout.  .Arch.  klin.  ^fcd.,  1016 
(110),  .')4S. 

i4d  See  Riven.  Arcli.  fjes.  Tliysiol..  1014    (157),  5S2. 


UlflV  ACID  !\    Till.    lil.Oiil).   77.S',S'L'£,S'  AND  URINE  627 

concent  i';it  ion  ot'  the  nrinc,  it  Ix'conu's  iii)i)ar('nt  liow  totally  devoid  of 
significant'C  is  the  presence  of  crystals  of  uric  acid  and  urates  in  the 
urine,  and  how  falUicious  is  any  theorization  based  upon  the  excretion 
of  considerable  quantities  of  uric  acid  when  all  the  above-mentioned 
factors,  especially  the  diet,  are  not  controlled  and  taken  into  consid- 
eration. Yet  on  just  such  an  inade(iuate  basis  has  been  constructed 
an  enormous  amount  of  theorization  as  to  "uric-acid  diathesis,"  "uric- 
acid  intoxication,"  "lithemia, "  etc.,  until  it  has  come  to  be  popularly 
believed  that  a  larg-e  share  of  the  minor  ailments  of  humanity,  and  in 
particular  all  non-infectious  diseases  of  the  joints  and  nuiscles,  are 
dependent  u])()n  the  presence  of  excessive  quantities  of  uric  acid  or 
urates  in  the  blood.  But  it  may  be  safely  stated  that  at  the  present 
time  there  exists  no  good  evidence  which  makes  it  probable  that  uric 
acid  is  responsible  for  any  pathological  conditions  whatever,  except 
uric-aeid  calculi,  "uric-acid  infarcts"  in  the  kidneys,  and  certain 
manifestations  of  gout.  Uric  acid  is  possessed  of  but  a  very  slight 
degree  of  toxicity,  and  the  body  is  able  to  get  rid  of  it  in  such  large 
measure  that  an  actual  intoxication  with  uric  acid  probably  never 
occurs. 

The  amount  present  in  the  urine  may  be  very  considerably  in- 
creased by  eating  food  rich  in  purines,  of  which  sweet-breads,  liver, 
and  kidney  are  the  best  examples ;  and  also  coffee  with  its  caffeine 
(trimethyl  purine),  may  give  rise  to  a  little  uric  acid,  although  the 
methylated  purines  seem  to  be  destroyed  in  large  part,  or  eliminated 
as  something  else  than  uric  acid.  Large  quantities  of  meat  will  also 
increase  the  uric  acid,  because  of  the  free  purines  contained  in  muscle ; 
and  even  a  diet  rich  in  proteins  free  from  purine  will  also  increase  the 
uric  acid  excretion  over  that  of  a  low  protein  diet.^"'  However,  the 
amount  of  uric  acid  in  the  blood  is  not  correspondingly  raised, ^'"^'^  this 
being  regulated  by  the  binding  function  of  the  tissues,  and  by  excre- 
tion through  the  kidneys.^^"  According  to  Folin  and  Denis  ^^  human 
blood  contains  1.5-2.5  mgs.  uric  acid  in  100  cc,  and  the  amount  bears 
no  fixt  relation  to  the  amount  of  urea  and  total  non-protein  nitrogen 
of  the  blood.^*"^  Any  difficulty  in  renal  elimination  is  usually  accom- 
panied by  an  increase  in  the  amount  of  uric  acid  in  the  blood,  in 
uremia  as  much  as  15  to  20  mg.  being  sometimes  found  per  100  c.c.^^" 
In  early  interstitial  nephritis  there  may  be  4  to  8  mg.  of  uric  acid  per 
100  c.e.  blood  without  a  corresponding  increase  in  urea  and  creatinine, 

15  Taylor  and  Rose,  Jour.  Biol.  Cliem.,  .1914    (18),  519. 

I'iaSce  Denis.  Jour.  Riol.  Chem.,   1915    (23),  147. 

15b  Stoeker  found  \nic.  acid  in  saliva,  increased  in  all  conditions  associated 
with  uricomia    ( Tnau^.  Dissert..  Zurich,  1913). 

ifi  Jour.  Biol.  C'heni.,  WU    (14).  29:   Arch.  Int.  Med.,  1915    (Ifi).  3:1. 

i'''.i  In  infants  tlic  amount  is  sliylith'  lower,  about  1.3  to  1.7  mu;.  Lii'finann. 
Zeit.   Kinderheilk.,   1915    (12).  227.' 

ir.b  See  Folin  and  Denis,  Arch.  Int.  :\rcd.,  1915  (16),  33;  :\revers  and  Fine, 
ibid.,  1916   (17),  570. 


628  URIC- ACID    METABOLISM    AND    GOUT 

which  suggests  tliat  uric  acid  may  be  less  easily  excreted  by  the  dis- 
eased kiduey  than  the  other  chief  nitrogenous  constituents  of  the 
urine. 

In  normal  individuals  there  seems  to  be  little  uric  acid  present  in 
the  tissues.  By  using  Folin's  method,  Fine  ^'"'  found  that  various 
tissues  contain  quantities  comparable  to  that  in  the  blood  of  the  same 
person,  whether  this  is  normal  or  increased  in  amount.  Ordinarily 
these  quantities  are  not  sufficient  to  permit  readily  of  isolation  of  the 
uric  acid  in  a  pure  state,  but  in  the  tissues  of  a  young  woman  who  died 
after  complete  suppression  of  urine  for  nine  days  following  poisoning 
with  HgCL,  I  found  considerable  amounts.^"''  AVhenever  much  de- 
.struction  of  the  nucleoproteins  of  the  tissues  is  occurring  in  the  body, 
the  elimination  of  endogenous  uric  acid  becomes  abnormally  raised,  the 
best  examples  being  the  resolution  of  pneumonic  exudates,  and  leu- 
kemia, especially  leukemia  under  .r-ray  treatment  {q.  v.).  In  neither 
of  these  conditions,  however,  do  any  symptoms  or  tissue  changes  arise 
that  can  be  referred  to  the  excessive  uric  acid. 


GOUT 

Introducing  this  subject,  one  cannot  do  better  than  to  quote  v. 
Noorden 's  statement  that  "It  is  not  to-day  very  alluring  to  write  am'- 
Ihing  regarding  the  theory  of  gout,  especially  in  a  book  which  is  essen^ 
tially  devoted  to  the  presentation  of  facts.  All  the  theories  advanced 
up  to  the  present  time  have  fared  badly.  The  positive  material  is 
much  too  insufficient  and  much  too  ambiguous."  After  adjusting 
the  many  contradictory  statements  of  earlier  investigators,  the  pres- 
ent status  of  our  conception  of  uric-acid  metabolism  in  gout  may  be 
briefly  summarized  as  follows :  The  excretion  of  uric  acid  in  patients 
with  chronic  gout,  when  kept  upon  a  definite  diet,  does  not  differ 
greatly  from  the  excretion  of  normal  individuals  on  the  same  diet. 
Normally  the  elimination  of  uric  acid  varies  within  rather  wide  limits, 
even  on  a  constant  diet,  but  the  excretion  in  chronic  gout  tends  to 
fall  at  or  slightly  below  the  lower  normal  limits.  As  a  rule,  gouty 
patients  on  a  purine-free  diet  excrete  less  endogenous  uric  acid  than 
normal  persons,  and  when  given  purines  in  the  food  the  rate  of  ex- 
cretion of  these  exogenous  purines  is  slower  than  normal.  According 
to  findzent's  studies,^^  in  nearly  all  cases  of  gout  the  blood  contains 
as  much  or  more  mono-sodium  urate  than  it  can  liold  in  solution  (8.3 
mg.  per  100  cc),  so  that  it  is  often  actually  a  supersaturated  solution 
of  tlie  relatively  insoluble  lactim  form  of  urate.  Even  on  a  purine- 
free  diet  the  blood  of  the  gouty  usually  contains  an  excess  of  uric  acid 
(4  to  f)  mg.  ]ier  100  cc),  independent  of  acute  attacks  or  any  marked 

inf.Tour.  KuA.  (')i.'iii.,  lt)l,-)  (2^),  47.3. 
i«'i.T(>ur.  Biol.  Cliom.,  lOlfi  (20),  ."^lO. 
IT  Zcit.  physiol.  Cliem.,  1909  (63),  4.55. 


GOUT  G29 

deficiency  in  the  cliniinatoi-y  jjowers  (jf  the  kidneys.  There  seems  to 
be  no  particular  relation  between  the  amount  of  uric  acid  in  the  blood 
and  the  occurrence  or  severity  of  attacks.'"''  This  uric  acid  is,  ac- 
cording: to  the  best  evidence,  in  a  free  state,  and  not  combined,  as  was 
at  one  time  urged  by  several  students  of  gout.  In  the  intervals  be- 
tween tlie  attacks  of  acute  gout  the  elimination  of  uric  acid  remains 
within  the  normal  limits;  however,  for  a  period  of  one  to  three  days 
before  each  acute  attack  the  amount  of  uric  acid  is  usually  decreased 
considerably.  With  the  onset  of  the  attack  the  amount  of  uric  acid 
excreted  becomes  increased,  and  for  a  few  days  remains  above  the 
average,  then  subsides  to  about  the  normal.  Of  these  two  features, 
the  increased  output  of  uric  acid  during  the  attack  seems  to  be  more 
constant  than  the  reduced  output  preceding  it,  but  cases  occur  in 
which  the  uric  acid  excretion  shows  no  variation  from  that  of  normal 
persons.  In  certain  cases  of  rheumatoid  arthritis  the  behavior  of  the 
purine  metabolism  resembles  that  of  gout.^* 

As  yet  we  have  no  definite  information  either  as  to  the  cause  of 
this  behavior  of  the  uric  acid  during  the  paroxysms  of  acute  gout, 
or  as  to  its  part  in  causing  the  paroxj^sm.  However,  in  view  of  the 
fact  that  monosodium  urate  is  found  in  the  joints  during  the  attacks, 
it  seems  most  probable  that  for  some  as  yet  unknown  reason  there 
occurs  a  precipitation  or  anchoring  of  the  urates  in  the  tissues,  which 
is  associated  with  the  attacks  of  pain  and  swelling.  We  do  not  know, 
however,  that  it  is  the  deposition  of  urates  that  causes  the  attacks. 
Indeed,  the  fact  that  uric-acid  retention  precedes  the  attack,  rather 
than  accompanies  it.  seems  to  suggest  that  it  is  the  absorption  of  the 
urate  rather  than  its  deposition  in  the  joints  that  is  responsible  for  the 
local  disturbances.  It  is  also  possible  that  during  the  period  of  re- 
tention the  uric  acid  is  held  in  the  blood  in  some  form  that  cannot 
be  eliminated  by  the  kidney,  and  that  its  deposition  in  the  joints  in 
an  absorbable  form  occurs  simultaneously  with  the  attack.  The  fail- 
ure of  recent  studies- on  the  enzymatic  transformation  of  purines  to 
locate  anywhere  in  the  human  body  an  enzyme  destroying  uric  acid, 
makes  hazardous  the  attempt  to  explain  gouty  metabolism  as  a  result 
of  enzymatic  abnormalities.  However,  there  can  be  little  doubt  that 
the  fundamental  reason  for  the  existence  of  uric  acid  gout  in  man  lies 
in  the  inability  of  the  human  organism  to  destroy  uric  acid.  Because 
man  cannot  destroy  uric  acid  rapidly  by  oxidation,  as  can  all  other 
mammals,  he  is  always  a  potential  victim  of  uric  acid  retention  and 
deposition. 

It  should  be  mentioned  in  addition  that  it  is  not  the  uric-acid 
metabolism  alone  that  is  altered  in  gout.     Irregular  periods  of  nitro- 

iTa  Pratt,  Amer.  Jour.  Med.  Sei.,  1916  (151),  92;  Bass  and  Herzherg.  Deut. 
Arch.  klin.  :\red.,   1916    (110).  482. 

18  W.  J.  Mallory.  .Tour.  Path,  and  Bact..  1910  (15),  207;  Ljungdahl,  Zeit.  klin. 
Med.,  1914   (49),  177. 


630  URIC-ACID    METABOLISM    AyD    (JOIT 

gen  retention  and  nitrogen  loss  are  quite  eonstant  features.  The 
cause  of  this  variability,  and  the  form  in  which  the  nitrogen  is  re- 
tained, are  quite  unknown,  although  there  is  some  evidence  that  the 
retained  nitrogen  is  in  the  form  of  purine  bodies  (Vogt).  Most  of 
the  excessive  loss  occurs  during  the  acute  attacks,^''  and  the  retention 
of  nitrogen  between  attacks  may  be  partly  to  repair  the  loss;  against 
this,  however,  is  the  fact  that  there  is  not  sufficient  gain  in  weight 
to  account  for  all  of  the  nitrogen  retention.  Associated  M'ith  the  de- 
layed excretion  of  ingested  purines  is  also  a  delayed  excretion  of  the 
other  nitrogenous  products  of  i)rotein  food.-"  The  proportion  of 
purine  bases  to  uric  acid  is  not  altered  in  gouty  urine. -^  The  state- 
ments in  regard  to  phosphoric  acid  elimination,  which  depends  largely 
on  decomposition  of  nucleins,  are  contradictory,  but  it  seems  probable 
that  it  shows  no  characteristic  alterations  in  gout.  Amino  acids,  espe- 
cially glycocoll,  are  said  to  be  excreted  in  excess.-^^ 

It  may  be  seen  from  the  foregoing  discussion  that  we  neither  under- 
stand fully  the  intricacies  of  metabolism  in  gout,  nor  know  whether 
uric  acid  is  responsible  for  either  the  acute  painful  attacks  or  for 
the  anatomical  alterations  in  the  kidneys,  heart,  and  bloodvessels. 
Indeed,  Daniels  and  McCrudden  -^^  have  shown  that  it  is  possible  for 
gouty  patients  to  have  a  persistently  low  content  of  uric  acid  in  the 
blood,  below  the  average  normal  quantity,  and  to  have  typical  acute 
attacks  without  change  in  either  the  uric  acid  content  of  the  blood  or 
its  excretion ;  attacks  were  even  observed  to  occur  when  the  blood  uric 
acid  was  at  a  subnormal  figure  from  administration  of  atophan,  which 
increases  its  elimination.  Furthermore,  Bass  and  Herzberg  -^"^  found 
that  uric  acid  can  be  injected  into  the  blood  of  gouty  subjects  until  the 
blood  contains  as  much  as  10  mg.  per  100  c.c.  without  causing  any 
joint  symptoms. 

It  is  very  possible  that  some  entirely  difiPerent  product  of  metabo- 
lism than  uric  acid  is  responsible  for  most  of  the  changes  and  symp- 
toms of  gout  -- — indeed,  this  would  seem  to  be  the  case  were  it  not  for 
the  great  frequency  of  the  deposition  of  monosodium  urate  in  the 
joints  and  cartilages,  both  during  the  acute  attacks  and  in  chronic 
gout.  This  indicates  that  there  is  surely  something  abnormal  in  the 
conditions  of  uric-acid  solution  and  circulation.  Why  the  urate  is 
precipitated  in  these  definite  places  is  another  of  the  many  unsolved 
problems  of  gout.  The  local  nature  of  the  deposition  indicates  that  it 
must  depend  upon  local  changes;  but  the  hypothesis  that  there  occur 

in  r{ruf,'scli,  Zcit.  cxp.  Patli.  u.  Tlicr.,  1000    (2),  filO. 
^"  I.ovcno  and   Kristeller,  .Tour.  Kxp.  Mod.,   1012    (Ki),   'MVA. 
ai  llcfrtor,  DiMit.  Arcl).  klin.  Mod.,  1!>1;{    (109),  :522. 
2i;i  IJiirtrcr  and  Scliwcririor,  Arcli.  oxp.  Tatli..   lOi:!    (74).  :?5;?. 
21b  Arcli.  Int.  Med.,  1!)].')    (l.'-i),  1040. 
•■:i<Deut.  Arch.  klin.  .Med.,  1010   (110),  482. 

--  In  swine  a  "<,nianinc  gout"  oceuris;  .see  Schittonlioini  and  Hendix,  /fit.  i)liv.-;iol. 
Ciioin.,  1000    (48),  140. 


GOUT  631 

first  (lt'<i-('iicrati\('  chaiigcs  in  the  tissues  wliit-h  deteniiiiKj  tlie  preeipi- 
tation  of  the  urate,  seems  to  have  been  disproved  by  the  demonstra- 
tion that  the  deposition  of  the  urates  precedes  the  necrosis.  The 
fact  that  the  presence  of  other  sodium  salts  in  a  solution  decreases 
the  solubility  of  urates  in  that  solution,  and  the  fact  that  cartilaf^e 
and  tendons  are  richer  in  sodium  salts  than  the  blood,  may  possibly 
have  something  to  do  with  the  fact  that  the  urates  are  precipitated  in 
these  particular  tissues.  On  the  other  hand  is  the  fact  that  in  leu- 
kemia and  nephritis  we  may  have  a  higher  concentration  of  uric  acid 
in  the  blood  than  in  gout,  and  this  urictemia  may  be  protracted,  with- 
out gouty  deposits  or  joint  symptoms.  Bass  and  Ilerzberg  -^''  found 
that  the  uric  acid  content  of  the  joint  fluid  was  approximately  the 
same  as  that  of  the  blood  in  patients  without  gout,  although  in  two 
gouty  uremics  they  found  18.5  and  20.8  mg.  in  the  joint  fluid  with 
only  10  and  8.2  mg.  in  the  blood.  They  also  found  that  intravenous 
uric  acid  injection  caused  less  uric^emia  in  the  gouty,  in  spite  of  re- 
duced renal  excretion,  and  hence  they  conclude  that  in  gout  the  tissues 
have  an  increased  capacity  for  taking  up  uric  acid. 

The  histology  of  urate  deposits,  both  experimental  and  gouty,  has 
been  carefully  studied  by  Freudweiler,^^  His,-*  Krause,-^  and  Rosen- 
bach.^''  Their  results  all  indicate  that  uric  acid  and  urates  excite 
some  slight  inflammatory  reaction,  cause  a  slight  local  necrosis,  and 
seem  to  act  as  a  weak  tissue  poison  (His).  However,  they  may  be 
deposited  without  causing  necrosis  (Rosenbach).  Possibly  part  of 
the  material  observed  in  areas  of  urate  deposition,  and  generally  con- 
sidered as  necrotic  tissue,  merely  represents  the  framework  of  the  crys- 
talline deposit  (Krause).  "When  experimentally  injected,  the  urates 
are  absorbed  slowly  by  phagocytic  leucocytes  and  giant-cells.  Why 
the  gouty  tophi  can  be  deposited  in  the  chronic  process  and  cause  no 
pain  or  inflammation,  while  in  acute  gout  deposition  of  urates  seems 
to  cause  such  marked  symptoms,  is  also  an  unanswered  question ;  un- 
less we  accept  the  explanation  that  the  slower  rate  of  deposition  and 
the  lack  of  dissolved  urates  account  for  the  absence  of  symptoms  with 
the  tophi.-'  ]\ragnus-Levy  holds  with  Pfeiff'er,  that  the  local  inflam- 
matory processes  must  be  ascribed  to  dissolved  urates,  since  they  often 
extend  for  some  distance  about  the  joints,  and  hence  the  attack  is 
ascribable  to  the  solution  rather  than  the  formation  of  the  deposits, 
a  fact  in  harmony  with  the  known  increased  elimination  of  uric  acid 
during  the  attack. 

That  urates  may  cause  necrosis  of  the  tissues  has  been  definitely 

23  Deut.  Aivh.  klin.  :\red..  ISnO    (r..3).  2li(). 

2i  Tbid.,  moo   (67),  81. 

25  Zeit.  klin.  Med.,  190.3    (50),  136. 

26VirdH>\v's   Arch.,    lOOo    (179).  359. 

2V  Almajria  (Hofnioistor's  Rcitr..  1905  (7),  466)  has  found  that  joint  cartilage 
]>laee<l  in  urate  solutions  becomes  fiUed  with  crystals,  which  infiltration  does  not 
occur  with  cartilage  of  anv  other  origin,  or  with  tendons. 


632  URIC-ACID    METAIiOl.lsU    AM)    GOUT 

established,  and  this  may  lead  to  conneetive-tissiie  formation  and 
contraction.-*  But  the  actual  increase  of  uric  acid  in  the  blood  and 
tissues  in  gout  is  so  slight  that  we  are  not  warranted  in  saying  that 
the  usual  tendency  to  sclerosis  in  all  the  organs  in  gout  is  due  to  the 
action  of  uric  acid,  rather  than  to  some  other  unknown  agent  or  agents. 
Excess  of  uric  acid  in  the  blood  is  by  no  means  pathognomonic  of  gout, 
for  we  may  have  relatively  great  excesses  of  uric  acid  in  the  blood  in 
leukemia,  in  some  cases  of  nephritis,  and  after  eating  large  amounts 
of  nucleoproteins,  without  a  symptom  of  gout.  Furthermore,  it  is 
quite  possible  that  the  precursors  of  uric  acid,  the  purine  bases,  are 
responsible  for  more  harm  than  the  uric  acid  itself.  Thus,  admin- 
istration of  adenine  to  dogs  and  rabbits  will  ])roduce  degenerative 
changes  in  the  kidneys,  associated  with  the  de])osition  of  substances 
resembling  uric  acid  and  urates  in  the  renal  tissue;  and  ]\Iandel  ^'^ 
states  that  purine  bases  may  cause  fever,  independent  of  infection. 
In  this  connection  it  may  be  mentioned  that  many  have  looked  upon 
renal  alterations,  leading  to  failure  of  excretion  of  uric  acid,  as  the 
primary  cause  of  gout ;  but  the  evidence  in  favor  of  this  is  faulty,  be- 
cause frequently  renal  changes  are  slight  or  entirely  absent  in  *gout, 
whereas  marked  nephritis  of  all  forms  may  exist  without  the  co- 
existence of  gout,  and,  as  mentioned  above,  the  kidney  in  gout  shows 
no  lack  of  ability  to  excrete  uric  acid  injected  into  the  tissues.  ]\Iag- 
nus-Levy,  however,  seems  to  believe  that  a  renal  retention  of  uric  acid 
is  of  importance,  and  that  it  may  occur  without  morphological  changes 
in  the  kidneys. 

The  newer  methods  of  blood  analysis  (Folin)  have  given  support 
to  this  view.  Fine  '^^  especially  calls  attention  to  the  fact  that  in 
early  interstitial  nephritis  the  blood  shows  a  greater  increase  in  uric 
acid  than  in  urea  or  creatinine,  as  if  the  diseased  kidney  found  more 
difficulty  in  excreting  uric  acid  than  the  other  substances.-^''  As  a 
result  the  blood  in  early  nephritis  may  show  quite  the  same  figures  for 
uric  acid,  urea  and  creatinine  as  are  found  characteristically  in  gout. 
Although  a  few  observers  have  occasionally  found  normal  amounts  of 
uric  acid  in  the  blood  in  gout,  this  seems  to  be  exceptional.  Hence 
we  are  still  confronted  with  the  question  whether  gout  is  anything 
more  than  a  form  of  nephritis  in  which  uric  acid  excretion  is  chiefly 
impaired,  or  whether  there  does  exist  a  special  disease,  gout,  which 
causes  uric-acidemia  more  or  less  independently  of  renal  abnormalities. 

2R  Ueeause  the  fjoiitv  toplii  do  not  siippurato,  even  wlicn  ulcoratod  tliroucrli  the 
skin,  it  has  boon  susrsrestod  that  the  urates  have  antisejitic  projierties.  'Bendix 
(Zeit.  klin.  IMed.,  1902  (44).  16.'i),  however,  could  not  donionstrate  such  antiseptic 
])roperties  experimentally.  Xo*  a1'"avs  do  tlie  tophi  consist  solely  or  even  lar<rely 
of  urate's,  hut  these  inav  he  replavcd  h\-  calciniii  salts  (  Kaliii.  Arcli.  Iiil.  ^led., 
ini3   (11),  92). 

2!>  Arncr.  Jour.  Phvsiol.,  1904   (10),  4r)2 ;   1907    (20),  4:?9. 
■'  2fiH  .Tour.   Anier.  :\Ied.   Assoc.   lOlH    ((JO),   20.')1. 
•  20b  See  also  Denis,  Jonr.  Hiol.  Chein..  191.5    (2.'?).   147. 


LA'/f  ACJD  JMWh'VTS  633 


URIC-ACID   INFARCTS  ■'■" 


Uric-aeitl  ini'arL-ts,  as  the  deposits  of  urates  and  urie  acid  observed 
in  the  kidneys  of  at  least  half  of  all  children  dying  within  the  first 
two  weeks  of  life  are  called,  ^ive  evidence  of  tlie  slijihtiiess  of  the  toxic 
effects  of  these  substances  upon  the  tissues.  Usually  little  or  no 
change  occurs  in  the  renal  tubules  as  a  result  of  these  depositions, 
except  such  as  can  be  attributed  to  their  mechanical  effect,^^  but  they 
may  serve  as  the  starting  point  of  calculi.  The  reason  for  the  for- 
mation of  these  infarcts  is  not  at  all  understood.  Spiegelberg  ^- 
found  it  possible  to  cause  them  experimentally  in  young  dogs,  in 
which  they  do  not  occur  naturally,  by  injection  of  0.25  gram  of  uric 
acid  per  kilo.  He  was  unable  to  explain  wh}^  this  deposition  should 
occur  in  young  animals  but  not  in  old,  for  he  could  not  find  evidence 
of  lessened  oxidative  power  on  the  part  of  young  animals,  and  the 
solvent  power  of  infants'  urine  was  found  equal  to  or  greater  than 
that  of  adults.  Other  authors,  however,  have  found  a  lower  oxidative 
power  in  young  animals,  and  ^Mendel  and  Mitchell  ^^  have  found  that 
in  the  embryo  pig  uricolytic  enzymes  do  not  appear  until  just  at  or 
just  after  the  time  of  birth.  As  human  tissues  have  no  demonstrable 
power  to  oxidize  uric  acid,  however,  these  animal  experiments  cannot 
be  applied  to  the  uric  acid  infarcts  in  human  infants.  Possibly  the 
uric-acid  infarcts  of  infants  are  the  result  of  the  great  destruction  of 
nucleoproteins  that  results  from  the  change  of  the  nucleated  fetal  red 
corpuscles  to  the  non-nucleated  adult  form.  Flensburg  believes  that 
a  hyaline  substance  is  secreted  in  the  urine  of  new-born  infants  which 
acts  as  a  matrix  for  urate  deposition.  jMcCrudden  considers  the  high 
concentration  of  infants'  urine  an  important  factor.  ^Minkowski  ^^ 
observed  that  administration  of  adenine  to  dogs  led  to  a  deposition  of 
uric  acid  or  some  similar  substance  in  the  kidneys.  Schittenhelra  ^^ 
found  the  same  deposits  in  the  kidneys  of  rabbits  fed  adenine,  but  not 
w^hen  they  w^re  fed  guanine.  According  to  Nicolaier.^"  the  crystals 
thus  deposited  are  not  uric  acid  or  urates,  but  6-amino-2-8-dioxy- 
purine,  derived  from  the  adenine  (6-amino-purine)  by  direct  but  in- 
complete oxidation.  He  could  not  find  this  substance  in  either  hu- 
man urine  or  in  a  uric-acid  calculus.     Eckert  ^'""■*  obtained  urate  de- 

30  See  discussion  hy  Wells  and  Corper,  .Tour.  Biol.  Cliein.,  1000   ((>).  .')21. 

31 1  have  obsej'ved  a  case  of  fatal  hematuria  veon-atonnti.  associated  with  most 
extensive  hemorrha<iric  infarction  of  both  kidricvs.  Tn  tlic  l)lnod\-  urine  B.  coli 
Avas  found  in  lar<re  nunilicrs.  From  the  anatomical  findin<rs  and  history  it  seemed 
quite  possil)lc  tliat  tlie  injury  of  tlie  kidneys  by  uric-acid  infarcts  mi£rht  have 
determined  the  localization  of  the  bacteria  in  these  orirans.  with  resulting 
hemorrliages.      (Trans.  Chicago  Path.  Soc.,   1000    (7),  242.) 

32Arch.  exp.  Path.  u.  Pharm.,  1808    (41),  428. 

33Amer.  Jour.  Phvsiol..  1007    (20),  07. 

3-t  Arch.  exp.  Path."  n.  Pharm.,  1808   (41),  .375. 

35/&UZ.,   1002    (47),  4.32. 

36Zeit.  klin.  Med.,  1002    (4.5),  350. 

36a  Arch.  exp.  Path.  u.  Pharm.,  1913    (74),  244. 


634  URICACID    METABOLISM    AXD    GOUT 

posits  by  intravenous  injection  into  ral)l)its  of  at  least  0.08  gm.  per 
kilo,  but  subcutaneous  injections  were  ineffective;  injury  to  the  renal 
epithelium  by  whatever  cause  interferes  with  this  deposition  of  urates. 
These  experimental  infarctions  are  undoubtedly  related  to  the  human 
form,  and  indicate  that  the  latter  dej)end  upon  the  presence  of  an  ex- 
cessive amount  of  uric  acid  in  the  infants'  urine,  in  which  a  ratio  of 
uric  acid  to  urea  of  7.9  to  74.9,  as  against  the  adult  ratio  of  about  2 
to  85,  was  found  by  Sjoquist.  According  to  Niemann,^^  in  .the  first 
few  days  of  life  the  infant  excretes  from  80  to  100  mg.  of  uric  acid 
daily,  while  after  the  fifth  day  the  amount  falls  to  30  to  40  mg.  daily. 
Similar  figures  were  obtained  by  Schloss  and  Crawford, ^^  who  also 
found  a  corresponding  increase  in  the  phosphoric  acid,  showing  that 
the  uric  acid  must  originate  from  nucleoproteins. 

Adult  kidneys  may  also  .show  uric  acid  deposits  in  the  tubules  of 
the  papilla?,  independent  of  gout.  They  occur  as  a  result  of  cell  de- 
composition, according  to  IM.  B.  Schmidt,^**  who  found  them  especially 
in  pneumonia,  leukemia  and  sarcoma,  but  not  in  carcinoma. 

37  Jahrb.  f.  Kinderheilk.,  1910  f71),  286. 
38Amer.  Jour.  Dis.  Cliildren.  1911    (1),  203. 
39  Verb.  dent.  Path.  Ges.,  1913   (16),  329. 


CHAPTER   XXI  I 

DIABETES 
By  R.  T.  Woody  ATT. 

Introduction. — As  witli  gout  and  the  problems  of  purine  metabo- 
lism, so  with  diabetes  a  vast  amount  of  study  has  been  expended  be- 
cause of  the  integral  couneetion  of  this  disease  with  the  broader  prob- 
lems of  carbohydrate  phj^siology,  the  metabolism  of  the  fats  and  the 
nature  of  internal  secretions.  It  is  impossible  in  this  place  to  review 
the  entire  literature  and  history  of  the  subject,  a  key  to  w'hich  will  be 
found  in  the  works  of  the  writers  cited  below. ^  This  chapter  will  be 
devoted  only  to  an  outline  of  the  chief  established  facts,  with  an 
indication  of  the  main  lines  along  which  the  thought  of  leading 
students  has  been  directed.  It  will  involve  a  brief  discussion  of  the 
problems  of  carbohydrate  pln'siology,  but  only  in  so  far  as  they  are 
contingent  upon  the  main  topic — while  for  a  discussion  of  the  anom- 

1  Older  Literature: 

Bouchardat — De  la  orlycosurie  ou  diabfete  sucre,  Paris,  1875. 
Kiilz — Beitriige  zur  Path,  und  Ther.  der  Diabetes  Melitus,  Marburg,  1874-5. 
Bernard — LeQons   sur   le    Dial)i'ti'   et   la   (Tlycogenese  Aniniale.    I'aris,    1877: 
Vorlesungen  iiber  Diabetes,  Berlin,  1878. 
Cantani — Diabetes  Melitus   (German  translation  by  Kalin),  Berlin,  1880. 
Frerichs — Ueber  den  Diabetes,  Berlin,   1884. 

Larger  Works: 

Naunyn — Der  Diabetes  ]\Ielitus,  Berlin,   1906.     Diabetes  Melitus;    in  Xoth- 

nagel's  Ilandbuch   (2nd),  Vienna,  1906. 
Lepine — Le  diabete  sucre,  Paris,  1909. 
von    Noorden — (a)     Die    Zuckerkrankheit    (6th),    Berlin,    1912.      (b)     Xew 

Aspects  of  Diabetes,  New  York,   1913. 
Pavy — Car))ohydrate  Metabolism  and  Diabetes,  London,  1906. 
McLeod — Diabetes    (Longmans,  Green),   1913. 

C'amniidge — Glycosuria  and  Allied  Conditions.      (Longmans,  Green),  1913. 
Allen — (jllvcosuria  and  Dial)etes,  Boston,   1913. 
Foster— Diabetes  .Melitus,  Philadelphia,   1915. 
Joslin — Treatment  of  Diabetes  Melitus,  New  York,   1916. 

Monographs,  etc. : 

Magnus-Levy — Diabetes  Melitus;  in  Kraus  and  Brugsch,  Spezielle  Path.  u. 
Ther.  innerer  Krankheiten,  Berlin,   1913. 

Gigon — Neuere  Diabetes  Forschungen.  Ergebnisse  der  inneren  Medizin, 
1912,  IX,  p.  206. 

von  Mering — Behandlung  der  Diabetes  melitus;  in  Penzoldt  and  Stinzing's 
Handbuoh  der  Spezielle  'I'herapie    1 2nd),    1912. 

Lusk — Elements  of  the  Science  of  Nutrition    (3rd),  New  York,   1917. 

Benedict  and  Joslin — Metabolism  in  Diabetes  Melitus,  Carnegie  Institu- 
tion, Washington,   1910. 

Benedict  and  Joslin — A  Study  of  ^letabolisni  in  Severe  Diabetes,  ibid.,  1912. 

635 


636  DIABETES 

alios  of  the  fat  metabolism  tlie  reader  is  referred  to  the  section  on 
aeidosis. 

Whereas  the  normal  urine  at  all  times  contains  reducing  sub- 
stances and  substances  Avliich  are  optically  active,  which  yield  crys- 
talline compounds  with  the  hydrazines  and  respond  to  other  so-called 
sugar  tests,  these  substances  are  not  all  sugars,  nor  are  all  the  sugars 
glucose.  The  quantity  of  fermentable  reducing  substance  in  normal 
urine  averages  about  4  parts  in  10,000  (0.04  per  cent.)  (Lavesson).- 
This  is  doubtless  subject  to  change  depending  upon  the  diet  and 
many  other  physiologic  factors.  The  total  reducing  substance  in  the 
normal  urine  of  adults  averages  0.21  to  0.24  per  cent.,  of  which  glucose 
constitutes  but  18  per  cent.  (Bang  and  Bohmannson).^  When  an 
abnormal  amount  of  sugar  occurs  in  the  urine  regardless  of  the  kind, 
the  condition  may  be  called,  in  accordance  with  Naunyn's  suggestion, 
melituria  (or  glycurui).  When  the  sugar  is  glucose  (dextrose),  the 
term  glycosuria  is  applied;  when  levulose,  Icvulosuria,  etc.  Other 
known  forms  of  melituria  are  lactosuria,  galactosuria,  fructosuria, 
pentosuria,  etc.  All  these  are  but  symptoms,  many  of  them  being 
caused  by  a  variety  of  mechanisms,  which  will  be  discussed  presently. 

The  term  diabetes  is  often  loosely  used  to  cover  any  variety  of 
inelituria,  but  it  is  preferably  limited  to  certain  forms ;  namely,  to 
the  glycosurias  (or  the  mixed  meliturias  in  which  d-glucose  is  the 
predominating  sugar),  and  further  than  this,  to  those  particular 
glycosurias  which  continue  even  after  the  glycogen  reserves  of  the 
body  have  become  depleted  and  when  the  diet  is  free  of  carbohy- 
drate ;  or,  to  those  transient  glycosurias  whose  nature  by  one  means  or 
another  can  be  proven  to  be  identical  with  the  continuous  forms  (la- 
tent or  mild  diabetes).  Over  against  these  are  the  meliturias  in  which 
other  sugars  than  glucose  play  the  chief  role,  and  glycosurias  which 
are  essentially  transient  because  they  depend  solely  on  the  ingestion 
or  administration  of  excessive  quantities  of  glucose  or  the  sudden 
liberation  into  the  blood  of  glucose  derived  from  preformed  glycogen 
or  other  fixed  compound  of  sugar. 

Thus,  the  gh-cosuria  which  follows  puncture  of  the  floor  of  the 
fourth  ventricle  (Claude  Bernard's  piqilre)  does  not  occur  in  animals 
which  contain  little  glycogen.  The  same  apj^lies  to  the  adrenal,  thy- 
roid and  hy])()pliysis  glycosurias.  But  after  comi)lete  pancreas  ex- 
tirpation (pancreas  diabetes)  and  in  the  spontaneous  human  disease 
(diabetes  melitus)  or  its  coujiterpai't  in  animals,  and  during  the  con- 
tinuous administration  of  ])lil()iliiziii,  the  glycogen  may  be  nearly  or 
(|uite  exhausted  and  the  diet  consist  solely  of  meat  and  fat  and  still 
the  glycosui'ia  continue.  On  the  other  hand  a  partial  ])ancreas  extir- 
pation, a  mild  diabetes  melitus,  oi-  an  interrupted  phlorhizinization 
may  give  rise  to  transient  glycosui-ia,  the  diagnosis  of  which  may  be 

zBioclii'iii.  /('it.,   1007    (4),  40. 

3  Zoit.  pliysiol.  Chem.,   1!)0;»,    (03),  443. 


CARHoii) Dh'ATh'  j'l/)  s/oi.oay  G37 

ilil'ticiilt.  Ill  ^'ciieral,  experience  teaches  tliat  all  persistent  j^lyco- 
surias  prove  to  be  diabetic  and  that,  except  in  i)liloi'hizin  poisoning;, 
every  fjemiine  diabetes  implies  a  disturbed  function  of  the  pancreas. 
In  forming  a  judgment  of  the  value  of  any  experimental  work  on 
diabetes  (histological,  chemical  or  clinical),  the  student  will  do  well  to 
examine  critically  the  records  of  quantitative  food  and  urinary  an- 
alyses offered  by  the  investigator,  to  show  what  type  and  what  grade 
of  diabetes  is  under  consideration. 

CARBOHYDRATE   PHYSIOLOGY 

Certain  facts  concerning  the  physiology  of  the  carbohydrates  may 
now  be  briefly  recalled  before  entering  into  the  discussion  of  the  in- 
dividual meliturias. 

The  appearance  of  sugar  in  the  urine  implies  a  source  or  sources 
of  sugar  and  the  existence  of  a  kidney  membrane  of  such  a  physical 
character  that  molecules  of  sugar  may  migrate  through  it  with  a 
certain  degree  of  facility.  The  factors  which  may  influence  the  purely 
physical  penetrability  of  the  kidney  membrane  to  sugar  molecules  are 
those  involved  in  a  discussion  of  kidney  function  and  secretion  in 
general  and  need  not  be  here  elaborated. 

Assuming  that  the  physical  penetrability  of  the  kidney  membrane 
to  sugar  molecules  is  normal  and  constant  there  are  then  two  basic 
moments  which  determine  how  such  sugar  will  pass  into  the  urine. 
These  are  1.  The  rate  at  which  sugar  molecules  enter  the  cells  consti- 
tuting tlie  kidney  membrane.  2.  The  rate  at  which  these  molecules 
of  sugar  undergo  chemical  change  into  something  else  within  the 
membrane. 

These  same  factors  of  supply  and  utilization  determine  the  elimina- 
tion of  sugar  from  any  cell  or  tissue  or  the  organism  as  a  whole,  but 
in  the  case  of  internally  situated  cells  the  elimination  is  directly  or 
indirectly  into  the  blood,  whereas  in  the  case  of  the  kidney  cells  sugar 
may  pass  into  the  urine  as  well  as  the  blood  and  thus  leave  the  body. 

Sugar  may  pass  out  of  a  cell  unchanged  when  the  rate  at  which 
it  enters  the  cell  from  internal  and  external  sources  exceeds  the  rate 
at  which  it  undergoes  change  into  something  else  within  the  cell,  the 
same  holding  in  the  case  of  the  cells  forming  the  kidney  membrane. 
Thus  sugar  passes  from  the  kidney  membrane  into  the  urine  when 
the  rate  at  which  sugar  molecules  enter  the  kidney  membrane  exceeds 
the  rate  at  which  they  are  denatured  or  utilized  wathin  it.  In  order 
to  understand  the  various  ways  in  which  melituria  may  be  produced 
it  is  necessary  to  analyze  farther  these  factors  of  supply  and  utiliza- 
iion. 

The  Sugar  Supply  to  the  Kidneys  is  chiefly  via  the  blood,  although 
the  metabolism  of  the  kidney  cells  may  lead  to  some  endogenous  sup- 
ply. The  factors  which  influence  the  rate  at  which  sugar  molecules 
enter  the  kidney  membrane  froiii  the  blood,  are  immedmte  and  remote. 


638  DIABETES 

The  immediate  factors  of  greatest  importance  are  the  blood  sugar 
concentration  and  the  area  of  kidney  membrane  exposed  to  the  blood. 
Even  a  casual  observation  of  the  kidney  meiii1)rane  as  it  appears  in  a 
glomerulus  with  its  capillary  loops,  capable  of  intlation  and  deflation 
with  varying  volumes  of  blood,  will  show  that  the  surface  of  contact 
between  the  blood  and  the  kidney  membrane  is  subject  to  wide  and 
easy  variations.  If  the  blood  sugar  concentration  remained  constant 
during  sucli  changes  of  the  expanse  of  membrane  in  contact  with  the 
blood  these  latter  changes  would  obviously  cause  variation  in  tlie  rate 
at  which  sugar  molecules  entered  the  membrane  from  the  blood.  Or, 
if  the  blood  sugar  percentage  varied,  the  rate  at  which  sugar  mole- 
cules entered  the  membrane  as  a  whole  might  remain  constant  if  the 
membrane  varied  its  expanse  in  direct  proportion  to  the  blood  volume 
and  in  inverse  proportion  to  the  blood  sugar  concentration.^  It  is 
therefore  apparent  that  observations  of  variations  of  the  blood  sugar 
concentration  can  not  afford  a  reliable  index  of  the  varying  rates  at 
Avhicli  sugar  enters  the  kidney  membrane,  unless  the  blood  volume,  or 
more  exactly  the  surface  of  contact  between  the  blood  and  cells,  is 
taken  into  account  and  the  same  principles  apply  in  the  case  of  all 
tissues  in  general.  Thus  Epstein  ^  observed  that  the  total  sugar  of 
the  blood  is  a  better  criterion  of  the  rate  of  glycosuria  in  diabetes 
than  the  blood  sugar  percentage.  The  more  remote  factors  which 
determine  the  quantity  of  sugar  brought  to  the  kidneys  via  the  blood 
are  the  supply  of  sugar  to  and  the  utilization  of  sugar  in  the  rest  of 
the  organs  of  the  body  apart  from  the  kidneys. 

The  Utilization  of  Sugar  may  be  considered  for  present  purposes  as 
the  sum  of  the  ])rocesses  by  which  a  sugar  such  as  glucose  is  converted 
into  something  else  within  the  cells.  In  the  case  of  glucose  it  includes 
oxidation  to  yield  finally  CO,  and  water;  poJ)jnierization  to  yield  a 
series  of  substances,  chief  of  Avhich  is  glycogen ;  reduction  to  fat ; 
transformation,  as  to  lactic  acid;  comhination,  etc.  The  rate  of  util- 
ization "  by  all  of  these  methods  taken  collectively  is  influenced  in  the 
first  place  by  the  rate  at  which  glucose  molecules  enter  the  cell  and 
secondly  by  the  reaction  conditions  encountered  within  it.  The 
utilization  rises  with  an  increasing  supply  of  glucose.  As  to  the  fac- 
tors which  enter  into  what  we  have  called  tlie  reaction  conditions 
found  within  the  cell  there  is  little  definite  knowledge.  With  a  con- 
stant glucose  sui)j)ly  the  rate  of  utilization  may  fall  as  the  result  of  a 

*  This  brings  up  tlio  question  of  the  froonietrical  forms  of  eapillaries.  It  is 
interesting  to  recall  tliat  if  a  hollow  cylinder  donbles  in  length  it  doubles  its 
volume  and  also  its  lateral  surface.  Tliis  tyjic  of  cajiiilary  distension  WMuld 
fulfill  tlie  above  conditions  and  other  methods  readilv  suggest  themselves. 

■•Jour.  Biol.  Chem.,  1014  (IS),  21:  Proc.  Roc.  Exp.  Biol..  lOlC  (1.3),  67:  also 
"Studies  in  Ilvperglvcemia  in  Relation  to  Clvcosuria,"  Albert  A.  Epstein,  X.  Y., 
1910. 

0  It  miglit  not  seem  desirabh'  (n  iiichidc  sucli  processes  as  temporary  storage 
in  the  form  of  glycogen  uiidcr  llic  licadiiig  i)f  utilization.  Tlie  term  is  used  for 
convenience. 


<•  {h'lKuivnh'ATi-:  I'll)  sKu.oav  6:v.) 

deficiency  of  that  liypotlictical  suhstancc  derived  iroiu  the  i)aiierea.s. 
It  is  well  known  tliat  acid  may  retard  glj^cogen  formation  and  hasten 
glycojren  hydrolysis.  It  would  appear  from  the  woi'k  of  Murlin  and 
Kramer"  that  alkali  may  inei'ease  jrlucose  utilization.  The  rate  of 
actual  oxidation  is  influenced  by  the  suj)i)ly  of  oxy<>:en,  etc.  The  fol- 
lowing may  serve  to  suggest  other  factors. 

It  mifiht  1)0  ooiu'oivcd  tliat  llu'  cell  coTilaiiicd  nioleciUcs  of  a  trhicolytic  catalyst 
or  enzyme  similar  in  its  effects  to  metallic  hydroxides,  that  jjlucose  molecules  as 
fast  as  they  entered  the  cells  would  come  into  collision  with  catalyst  molecules, 
perhaps  combining  witli  tliem,  and  that  as  a  result  of  the  encounter  the  glucose 
molecules  would  he  dissociated  into  unsaturated  frajjments  or  ions.  From  the  mo- 
ment of  imion  or  dissociation  they  would  cease  to  behave  as  glucose  molecules. 
Tlie  unsaturated  fragments  might  suhs('(|ueiitly  sutler  various  fates,  depending 
ujion  tlie  cliaracter  and  quantities  of  various  substances  in  tlie  cell.  Thus,  some 
might  combine  with  oxygen  to  yield,  finally,  carbon  dioxide  and  water.  Others 
might  combine  with  each  other  to  form  polymers  like  glycogen,  others  again 
undergo  reduction  to  fat  or  molecular  rearrangement  to  give  lactic  acid.  The 
relative  quantities  imdergoing  those  several  changes,  would  depend  upon  the 
relative  quantities  of  H  and  OH  ions,  of  available  oxygen,  salts,  etc.,  foimd  in  the 
various  phases  of  the  cell.  This  conception  is  based  on  that  used  by  Nef  to  ex- 
plain the  behavior  of  sugars  in  alkaline  solutions.  For  a  concrete  conception  of 
the  dvnamics  of  a  reaction  between  an  organic  substrate  and  catalyst  the  reader 
is  referred  to  Van  Slyke's  study  of  the  enzyme  urease.s 

The  general  principles  outlined  above  may  be  illustrated  by  experi- 
ments with  timed  intravenous  injections  of  glucose.  It  has  long  been 
known  that  if  a  comjiaratively  large  dose  of  glucose  is  injected  rapidly 
into  a  peripheral  vein  a  marked  glycosuria  usually  results.  Pavy, 
however,  emphasized  the  fact  that  a  material  fraction  of  a  dose  so 
given  fails  to  be  excreted  and  appears  to  be  utilized.  Doyon  and 
Du  Fourt  demonstrated  that  with  a  standard  dose  of  gluco.se  the  per- 
centages excreted  and  utilized  respectively  are  influenced  by  the  time 
consumed  in  injection,  the  slower  rates  of  injection  causing  lower  per- 
centage excretions  and  vice  versa.  Blumenthal  chose  a  standard  in- 
jection time  of  about  10  seconds  and  varied  the  weight  of  sugar  given 
in  that  time.  He  found  that  a  certain  dose  of  glucose  might  be 
injected  into  the  ear  vein  of  a  rabbit  without  causing  any  glycosuria 
at  all.  However,  the  maximum  dose  which  could  be  so  injected  once 
could  not  be  repeated  15  minutes  later  without  causing  glycosuria. 
He  assumed  from  this  that  the  first  dose  "sattirated"  the  tissues  and 
that  fifteen  minutes  later  the  utilization  of  sugar  had  only  resulted  in 
a  partial  desaturation.  He,  therefore,  determined  the  dose  of  glucose 
which  might  be  injected  repeatedly  at  15  minute  intervals  for  as  long 
as  3  hours  without  ever  causing  glycosuria.  His  figures  varied  be- 
tween 0.6  and  1.3  gm.  per  kg.  of  body  weight  per  hour.  This  he 
termed  the  "utilization  limit,"  whereas  the  largest  dose  which  could 
be  given  within  10  seconds  once  without  causing  glycosuria  he  called 
the  "saturation  limit."     The  latter  he  placed  at  0.8  gm.  per  kg.  but 

7  Jour.  Biol.  Chem.,  101  fi   ^27),  490. 

8  Jour.  Biol.  Chem.,  1014   (10),  141. 


640  DIABETES 

E.  ]\r.  Wilder  has  been  unable  to  confirm  this  observation.  Woodyatt, 
Sansum  and  Wilder  ^  made  continued  intravenous  injections  of  glu- 
cose at  uniform  rates  by  means  of  a  motor  driven  pump  for  2  to  17 
hours  with  the  following-  findings : 

If  chemically  pure  glucose  in  aqueous  solution  is  injected  con- 
tinuously into  the  peripheral  venous  blood  of  a  normal  resting  man, 
dog  or  rabbit  at  the  rate  of  0.8  gm.  per  kg.  of  body  weight  per  hour, 
or  at  any  slower  rate,  the  injection  may  be  sustained  in  most  cases, 
hour  after  hour  for  7  hours  and  probably  longer  without  producing 
any  glycosuria  in  the  usual  sense  of  the  word.  If  the  rate  is  advanced 
to  0.9  gm.  of  glucose  per  kg.  of  body  weight  per  hour,  while  all  other 
conditions  remain  the  same,  the  injection  may  be  sustained  for  a  short 
time  without  causing  glycosuria,  but  in  nearly  all  cases  abnormal 
quantities  of  glucose  beg^in  to  appear  in  the  urine  after  5  to  30  minutes 
of  injection,  the  time  depending  upon  the  previous  degree  of  satura- 
tion. Once  established,-  the  glycosuria  then  tends  to  proceed  at  a 
uniform  rate  as  long  as  the  rate  of  injection  and  other  conditions  re- 
main fixed.  However,  if  the  injection  rate  is  again  reduced  to  0.8  gm. 
per  kg.,  glycosuria  promptly  ceases.  Thus  during  the  continuance  of 
an  injection  at  the  latter  (0.8  gm.)  rate  there  can  be  no  continued 
accumulation  of  unchanged  glucose  in  the  body,  but  the  rate  of  injec- 
tion is  equalled  hy  the  rate  of  utilization.  It  is  important  to  note 
that  it  makes  no  appreciable  difference  whether  one  uses  an  18  or  a 
72  per  cent,  glucose  solution  for  injection.  The  tolerance  limit  for 
glucose  may  be  demonstrated  at  the  same  point  regardless  of  wide 
variation  in  the  quantity  of  water  administered  with  the  glucose,  even 
1  hough  the  blood  volume  and  the  blood  sugar  percentages  may  be 
influenced  by  variation  of  the  water  supply.  Also,  if  glucose  is  in- 
jected continuously  and  uniformly  at  a  rate  productive  of  some 
glycosuria,  the  glucose  excretion  may  proceed  at  a  constant  rate  in 
spite  of  marked  variation  in  the  water  supply  during  successive  hours. 
A  certain  dog  receiving  by  vein  20  gm.  of  glucose  per  10  kg.  per  hour 
for  8  hours,  excreted  every  hour  close  to  0.42  gm.  of  sugar  per  10 
kg.  of  body  weight.  Yet  during  the  experiment  water  was  injected 
at  varying  rates  into  the  same  vein  with  the  glucose,  so  that  the  hourly 
volume  of  urine  varied  between  6  c.c.  and  128  c.c.  and  the  percentages 
of  sugar  in  the  ui'ine  varied*betvveen  0.35  and  4.9.  This  em])hasizes 
the  fundamental  importance  of  the  rate  at  which  sugar  is  supplied  to 
the  organism  in  determining  the  occurrence  or  non-occurrence  of 
glycosuria  and  in  fixing  the  rate  of  excretion  when  the  latter  occurs. 

In  view  of  the  above  generalities  several  specific  mechanisms  sug- 
gest themselves  by  which  glycosuria  might  be  produced  : 

(1)  An  increased  supply  of  preformed  glucose  to  the  whole  organ- 
ism from  without  (alimentary  glycosuria). 

f  Jour.  Amor.  M(>d.  Assoc,  101.')  ((>")).  2007  (preliminary  report);  Woodyatt, 
ITarvev  Sociptv  Loftnros,  IDIO;  Wilder  and  Sansnm,  Arch.  Int.  Med..  1017  (10), 
311;  Woodyatt  and  Sansiun,  Jour.  Biol.  Ciiem.,  1017    (30),  155. 


77//;  iii.(K)h  srcM!  641 

(2)  A  (Iccrcascd  utilization  in  tlie  orgjaiiism  as  a  whole  (j)aiicreatic 
diabetes). 

(8)  All  iiit-reased  supply  to  tlie  kidneys  resultinj^  from  the  libera- 
tion into  the  blood  of  sugar  previously  stored  or  combined  in  other 
orp-ans.  Thus,  the  rapid  hydrolysis  of  glycogen  following  ])un('ture 
of  the  floor  of  the  fourth  ventricle  and  analogous  nerve  stiimilations, 
and  occurring  in  the  acid,  asphyxial,  narcotic,  thyroid,  epinephrine, 
and  hypophysis  glycosurias.  In  an  analogous  manner  lactose  may 
enter  the  circulating  blood  from  the  mammary  gland,  and  pentose 
from  unknown  sources. 

(4)  An  increased  supply  to  the  kidneys  due  to  decrea,sed  utilization 
in  other  organs.  The  breaking  down  of  glycogen  mentioned  in  (3) 
might  be  so  interpreted. 

(5)  Decreased  utilization  in  the  kidney  itself. 

(6)  Increased  physical  penetrability  of  the  kidney  membrane  to 
glucose.  Both  (5)  and  (6)  are  hypothetical  conditions,  the  latter 
having  been  proposed  as  the  basis  of  so  called  kidney  diabetes,  a  state 
in  which  glycosuria  occurs  with  a  normal  or  subnormal  percent  age 
of  sugar  in  the  blood  and  in  which  the  rate  of  sugar  excretion  is,  in 
comparison  with  other  forms  of  glycosuria,  little  influenced  by  the 
diet.-"^ 

THE  BLOOD  SUGAR  * 

The  normal  blood  sugar  concentration  is  found  to  average  0.10 
per  cent.,  but,  as  statistics  show,  it  may  vary  at  least  between  0.06  and 
0.11  per  cent.  The  literature  contains  numerous  references  to  that 
blood  sugar  concentration  which  if  exceeded  leads  to  glycosuria 
("threshhold"  value).  In  accox'dance  with  the  general  principles 
above  discussed  we  should  expect  this  value  to  vary.  It  has  been 
placed  at  0.147  to  0.164  per  cent,  by  Foster,  between  0.17  and  0.18 
per  cent,  by  Hainan  and  Hirschman,  at  about  0.20  per  cent,  by  Pavy, 
and  other  writers  have  reported  greater  variations,  due  in  part  doubt- 
less to  differences  in  the  analytical  methods  used.  How  widely  the 
threshhold  blood  sugar  percentage  may  be  varied  by  extreme  variations 
of  the  blood  volume  and  other  factors  has  not  been  settled.  Following 
the  ingestion  of  free  glucose  the  blood  sugar  percentage  ordinarily 
rises,  and  in  a  similar  way,  but  more  slowly,  after  feedings  of  starch. 
Fisher  and  "Wishart  gave  50  gm.  of  glucose  in  150  c.c.  of  water  by 
stomach  to  dogs  weighing  8  to  0  kg.  and  found  in  the  first  hour  blood 
sugar  percentages  of  0.16  and  0.13.  In  succeeding  hours  there  was 
little  variation  from  0.11  per  cent.  In  harmony  Avith  the  previous 
work  of  Gilbert  and  Baudoin  and  the  more  recent  studies  of  others  on 
man.  these  experiments  showed  that  the  l)lood  sugar  percentage  rises 
during  the  first  hour,  then  falls  and  thereafter  remains  iKU-mal.  There 
was  no  increase  of  the  blood  volume  during  the  first  hour,  the  hemo- 

9a  Cf.  Epstein,  A.  A.  ^rosenthal. 
41 


642  DIA  BETES 

globiu  pereeiitajio  remaiiiinp'  iiiiehaiigod,  jn-obably  because  the  large 
quantity  of  ghieose  in  the  bowel  held  water  there.  But  in  the  second 
hour  the  blood  volume  became  large  and  the  hemoglobin  showed  the 
effects  of  dilution.  In  this  same  hour  the  sugar  percentage  returned 
to  normal.  But  the  absorption  of  glucose  was  only  completed  in  the 
fourth  lumr  and  calorinietric  observations  by  Lusk  showed  that  the 
metabolism  also  ran  at  a  uniform  rate  20  per  cent,  above  the  basal 
level  into  the  fourth  hour.  Accordingly  the  observed  blood  sugar  per- 
centages first  rose  as  tlie  rate  of  sugar  supply  was  increased,  but  fell 
again  during  the  )i\ainte)iance  of  the  increased  siipphj  and  ichih  the 
metabolism  was  constant,  owing  to  the  shifting  of  water. 

When  concentrated  (54  to  72  per  cent.)  glucose  solutions  are  in- 
jected continuously  into  the  blood  at  rates  of  0.4  to  0.8  gm.  per  kg. 
per  hour,  there  is  at  first  a  steep  rise  of  the  blood  sugar  percentage, 
followed  by  a  fall  coincident  with  an  increased  hydremia,  after  whicli 
a  new  equilibrium  is  established  and  the  blood  sugar  percentage  may 
become  constant  at  a  "normal"  level  exactly  as  in  the  above.  By 
injecting  glucose  at  the  same  rates  in  sufficiently  dilute  solutions  this 
initial  rise  may  be  very  much  reduced  and  the  ])lood  sugar  percentage 
established  in  later  hours  may  even  be  lower  than  that  observed  before 
injection  began.  On  the  other  hand,  if  glucose  is  injected  at  rates 
above  0.9  gm.  per  kg.  per  hour,  glycosuria  begins,  and  if  the  rate  of 
injection  is  rapid  enough  may  be  made  intense.  As  glucose  passes 
through  the  kidney  membrane,  water  tends  to  accumulate  with  the 
glucose  on  the  urinary  side  of  the  membrane  (increased  diuresis, 
polyuria ).  In  the  same  way  that  glucose  in  the  bowel  lumen  may  tend 
to  withhold  water  from  the  blood,  so  a  sufficient  quantity  of  glucose 
in  the  urinary  tubules  may  manifest  the  same  tendency  in  this  local- 
ity. Whether  the  glucose  in  the  urinary  tubules  will  have  the  effect 
of  concentrating  the  blood  or  vice  versa  will  depend  on  the  cpiantitative 
distribution  of  free  sugar  between  tliese  two  fluids,  and  the  ((uantity 
of  water  available  for  distribution  between  the  blood  sugar  and  the 
urinary  sugar.  During  continuous  intravenous  injections  of  glucose 
at  rates  from  2.7  gm.  per  kg.  per  hour  upward.  80  to  40  per  cent, 
of  the  glucose  injected  may  be  excreted  and  there  is  a  strong  tendency 
toward  dehydration  of  the  whole  body.  This  may  be  neutralized  b\" 
su])plying  water  with  the  sugar  as  fast  as  it  flows  away  in  the  urine, 
provided  the  rate  of  injection  is  not  so  great  that  tlie  necessary  traffic 
in  water  overtaxes  the  cardio-i-enal  mechanism.  By  employing  these 
high  rates  of  injections  and  maintaining  the  water  balance  at  as  low 
levels  as  c(mipatil)le  with  life  and  recovery,  it  is  possible  to  produce 
and  maintain  for  houi's  blood  sugar  concenti'ations  as  higli  as  2.;?8  ]K'r 
cent.  Joslin  obsei-ved  1.40  ])er  cent,  of  sugai'  in  llic  blood  of  a  fatal 
case  of  diabetes  with  nephi'itis.  This  is  ])rol)ably  the  highest  on 
record.     Tlie  blood  sugai-  of  diabetics  passing  sugar  in  the  ni'ine  is  as 


77//;  N7M77-;  OF  Till:  srcM!  i\  Tin:  r.i.ooh  643 

a  rule  hiiilicr  tliaii  iiDniial.  l)iit  not  iicct'ssafily  so,  iiiucli  (l('])eM(liii<z;  on 
the  watci'  halaiu't'.  .loslin's  statistics  show  a  raii^e  of  0.07  to  0.4:> 
per  cent. 

THE  STATE  OF  THE  SUGAR  IN  THE  BLOOD 

It  has  k)n<>-  been  believed  that  tlie  sii<iar  circiUatin<;'  in  the  Ijlood 
exists  ill  two  physical  states,  a  diffusible  and  a  non-difrnsiblc,  i.e.,  as 
(a)  Free  ghicose  in  a  state  comparable  to  that  of  glucose  dissolved  in 
water,  sucre  actueUe  of  Lcpine.  (b)  Sugar  in  a  colloid  state,  Sucre 
rirtuelle,  Lepine.  By  the  former  term  a  very  specific  idea  is  conveyed. 
One  might  think  for  instance  of  single  molecules  or  clusters  of  two  or 
three,  each  holding  in  its  sj^here  of  intluence  a  certain  number  of 
water  molecules  like  suns  in  solar  systems.  Such  small  masses  move  at 
high  velocities,  "diffuse"  readily  and  create  high  "osmotic  pres 
sures. "  By  the  latter  term  is  meant  sugar  in  the  blood  which  does 
not  behave  physically  like  glucose  in  aqueous  solution  nor  respond  to 
the  ordinary  chemical  tests  for  sugar,  but  from  which  free  glucose  may 
be  reliberated  by  such  simple  procedures  as  boiling  with  dilute  acids. 
Such  sugar  is  supposed  to  exist  as  a  component  of  particles  having  the 
larger  dimensions  which  characterize  colloids  (non-diffusoids).  But 
as  to  the  actual  chemical  nature  of  these  a  great  variety  of  proposals 
have  been  made.  Thus  Pavy  proposed  glucose  molecules  held  entire 
to  the  colloids  of  the  blood  in  a  state  of  simple  adsorption  (comparable 
to  the  state  of  molecules  of  a  dye  electrically  bound  to  particles  of  a 
colloid  clearing  agent).  He  also  proposed  glucose  chemically  incor- 
porated in  the  structure  of  the  protein  molecule,  and  between  these 
extremes  by  the  same  author  a  score  of  suppositions  have  been  made 
by  others,  among  which  Drechsel's  jecorin,  a  lecithin-sugar  compound, 
is  a  notable  example.  Another  worthy  of  serious  consideration  is 
that  of  glucose  built  up  into  polymers  intermediate  between  disac- 
charides  and  glycogen.  The  basis  for  assuming  the  existence  of  com- 
bined sugar  in  the  blood  lies  chiefly  in  the  observation  that  following 
glucose  administrations  the  increase  in  the  reducing  power  of  the 
blood  which  results  from  heating  the  blood  with  dilute  acid  is  greater 
than  the  increase  resulting  from  the  same  process  before  sugar  admin- 
istration (see  Pavy,  Lepine,  Loewi).  Also,  if  glucose  is  added  to  fresh 
blood  and  the  mixture  placed  in  the  incubator,  the  reducing 
power  falls  but  may  be  in  part  rehabilitated  by  boiling  with  dilute 
acid.^" 

As  to  the  sugar  wliidi  is  determined  by  the  ordiiuiry  methods 
of  blood  analysis,  it  would  ai^jieai-  that  we  are  dealing  almost  ex- 
clusively with  free  glucose.     As  yet  no  one  has  succeeded  in  prov- 

10  A  critical  review  of  the  literature  to  1012  will  he  found  in  the  hooks  hy 
IMcLeod  and  Allen.  Compare  also  the  article  hv  Levene  and  the  recent  studies 
of  Lombroso  favoring  the  polymerization  idea. 


644  DIABETES 

jn<i"  the  existence  in  blood  of  a  combined  sutr'ar  capable  of  spontaneous 
conversion  into  free  suoar.  Michaelis  and  Rona  dialyzed  separate  por- 
tions of  the  same  blood  against  isotonic  salt  solutions  containing 
graduated  quantities  of  sugar.  A  sugar  solution  which  neither  lost 
nor  gained  sugar  during  the  dialysis  they  regarded  as  having  an 
amount  of  free  sugar  e((ual  to  that  in  the  blood.  Titration  of  the 
blood  sugar  and  of  the  sugar  in  such  a  solution  gave  almost  identical 
figures.  They  accordingly  concluded  that  all  of  the  reducing  sugar 
in  this  blood  must  have  been  as  free  to  diffuse  as  was  that  in  the  simple 
salt  solution.  But  this  ingenious  experiment  of  ^lichaelis  and  Rona 
does  not  show  conclusively  that  in  circulating  blood  there  is  no  sugar 
in  a  state  of  colloidal  adsorption,  because  drawn  blood  rapidly  under- 
goes survival  changes  (e.g.,  lactic  acid  formation)  which  might  influ- 
ence the  affinity  of  its  colloids  for  sugar.  However,  McGuigan  and 
Hess  ^^  led  the  blood  of  living  animals  thi-ough  collodion  tu])es  en- 
closed in  jackets  filled  with  isotonic  salt  solutions  and  found  that  when 
equilibrium  was  established  the  concentration  of  reducing  sugar  in 
the  salt  solution  and  in  the  plasma  was  the  same,  proving  that  even 
in  life  all  of  the  titrable  plasma  sugar  is  in  a  state  of  subdivision  which 
lets  it  migrate  through  the  interstices  of  a  collodion  membrane.  This 
would  make  the  adsorption  idea  seem  untenable. 

Closely  related  to  the  question  of  the  state  of  the  sugar  in  the  blood 
is  that  of  its  state  in  the  cells.  Palmer  ^-  has  studied  the  percentages 
of  sugar  found  in  the  various  tissues  in  relationship  to  the  plasma 
sugar  concentration.  The  titrable  sugar  of  the  tissues  was  found 
below  that  of  the  blood  in  all  organs  except  the  liver.  Of  course, 
owing  to  the  large  quantities  of  glycogen  which  occur  in  that  organ 
and  the  rapidity  with  which  it  breaks  down  into  glucose,  liver  tissue 
would  naturally  analyze  high  for  sugar.  In  the  muscles  the  titrable 
sugar  was  found  at  0.04  and  0.041  per  cent,  with  blood  sugar  at  0.10 
and  0.105  per  cent.  On  the  other  hand  the  tissues  generally  when 
boiled  with  dilute  acid  show  a  higher  content  of  "combined"  sugar 
than  the  blood.  Tliis  is  most  striking  in  the  case  of  the  liver  and 
due  by  common  consent  to  the  polymers  of  glucose  in  that  organ. 

It  serves  a  useful  purpose  to  consider  the  body  as  a  whole  as  an 
heterogeneous  system  made  up  of  phases,  and  to  assume  that  glucose 
on  entering  the  body  distributes  itself  between  these  phases  as  acetic 
acid  may  distribute  itself  between  the  fat  droplets  and  the  acpieous 
part  of  milk ;  that  glucose  in  a  certain  type  of  phase  behaves  as  though 
in  water  and  in  anothei-  type  of  phase  rapidly  undergoes  chemical 
changes.  The  blood  is  a  tissue  in  which  the  dominant  phase  is  in  the 
iialui-e  of  a  ])hysical  solvent  foi"  glucose,  like  water.  In  the  cells  the 
dominant  phases  are  of  sucli  a  character  that  glucose  on  entering  Ihcm 

11  Jour.  Pliarin.  and  Exp.  Tlior.,    (1014)     (G),  45. 

12  Jour.  Biol,  (hem.,  1917   (30),  79. 


THE  HT.VVK  OF  Tin:  Hl(!.\h'  l\    Till:  HLDOh  645 

rai)icll\-  uiuler<iULS  eiiemical  cluinge.  But  botli  types  of  phase  are 
present  in  both  blood  and  cells  although  in  different  proportions. 
The  blood  is  therefore  the  phase  par  excellence  in  which  to  study  the 
" Sucre  actuelle"  and  the  tissues  the  place  to  study  the  "sucre  vir- 
tuelle."  According  to  this  conception  "sucre  virtuelle"  would  be 
glucose  in  process  of  utilization  or  storage  and  not  beyond  recall, 
hence  chiefly  glucose  polymers. 

DIOSE  13 

Diose,  glyt'ollic  aldeliyde,  CH.OH-COH,  the  simplest  sugar,  of  whicli  there  is 
but  one  possible  form,  is  higlily  sensitive  to  oxidative  infhiences  and,  in  vitro, 
readily  roiidenses  with  alkali  to  yield  a  complex  mixture  of  higher  sugars  and 
saccharinic  acids  in  a  manner  analogous  to  that  manifested  liy  tlie  trioses.  Not- 
withstanding its  instability  and  sensitiveness  to  oxidative  clianges  in  the  test 
tube,  it  would  appear  that  glycollic  aldehyde  is  insusceptible  of  direct  oxidation 
in  the  body  but  that  it  may  be  converted  into  glucose,  like  other  sugars,  and  then 
utilized.  When  given  intravenously  at  the  rate  of  only  0.1  gm.  per  kg.  per  hour, 
unchanged  diose  appears  in  the  urine  after  the  first  few  minutes  of  injection 
(author).  P.  Mayer  reported  glycosuria  and  death  following  administration  of 
imi)ure  glycollic  aldehyde  to  rabbits.  Parnas  and  Baer  saw  an  increase  of  glycogen 
in  tortoise  livers  perfused  with  glycollic  aldehyde.  This  is  confirmed  by  Barren- 
scheen.  Smedley  noted  the  rapid  disappearance  of  diose  added  to  liver  emulsions. 
Sansum  and  Woodyatt,  and  also  Greenwald  observed  slight  increases  of  the 
glycosuria  following  parenteral  administrations  of  diose  in  phlorhizinized  but  not 
completely  deglycogenized  dogs.  The  extra  sugar  could  liave  come  from  glycogen 
in  these  experiments.  A  Hnal  proof  of  the  conversion  of  diose  into  glucose  in  the 
living  bodv  has  not  been  brought.  The  relationship  of  this  substance  to  glvcine, 
CH.XH,-COOH;  glycollic  acid,  CH,OH-COOH;  and  ethyl  alcohol,  CH.rCH.OH;  is 
close.  Lusk  states  that  glycine  is  capable  of  conversion  into  glucose  in  the  body. 
However,  glycollic  acid  and  alcohol  are  apparently  not  sugar  formers. 

TRIOSES  11 

There  are  three  possible  trioses,  d-  and  1-glyceric  aldehyde  and  the  ketotriose 
dihydroxyacetone.  Tlie  optically  inactive  d,  1-glyceric  aldehyde  has  been  jircpared 
ami  recently  the  d-  and  1-forms.  The  preparation  is  still  tedious  and  expensive. 
Dihydroxy-acetone  is  somewhat  easier  to  prepare.  Both  trioses  are  imstable, 
easily  oxidized  and  very  prone  to  undergo  rearrangements  and  condensation  with 
even  traces  of  alkali.  lender  the  inlluence  of  alkali  they  yield  complex  mixtures  of 
hexoses,  chiefly  S-kctohexoses,  formerly  known  as  a  and  /3-acrose  from  which 
Schmitzis  has  recently  isolated  d,l-fructose  and  d,l-sorbose.  If  oxygen  is  avail- 
able as  well  as  alkali,  they  burn.  If  the  alkali  is  strong  and  oxygen  lacking,  much 
lactic  acid  is  formed  together  with  certsiin  rearranged  tetrose,  pentose  and  hexose 
molecules,  known  as  saccharinic  acids  (or  "saccharines,"  of  Kiliani).  The  same 
phenomena  occur  when  the  alkali  is  dilute,  but  more  slowly.  The  structural 
formuht  of  the  trioses  and  their  relationship  to  glycerol,  glyceric  acid  and  lactic 
acids  (the  latter  of  which  might  be  regarded  as  a  .3-carbon  saccharinic  acid)  may 
be  seen  from  the  following  chart: 

13  Literature  on  diose:  Mayer,  P.,  Zeit.  f.  physiol.  Chem.,  100,3  (.38).  135; 
Woodvatt,  R.  T.,  .Tour.  Amer.'  Med.  Assoc,  1010 '(5.5).  2100;  Parnas  and  Baer, 
Biochem.  Zeit.,  1012  (41),  386;  Smedlev,  Ida,  .Tour.  Phvsiol.,  1012  (44),  203; 
Sansum,  W.  D.  and  Woodyatt,  R.  T.,  .Toiir.  Biol.  Chem.,  1014    (17)   r)21. 

1*  Literature  on  Trioses:  The  cliemical  literature  is  reviewed  and  an  improved 
method  of  preparing  glyceric  aldehyde  described  by  Witzemann,  K.  .!.,  .Ttnir.  Am. 
Chem.  Soc,  1014  (36)",  lOOS,  and  ihid.,  p.  222.3.  The  biological  literature  is 
reviewed  bv  Sansum,  W.  D.  and  Woodvatt.  15.  T..  .Tour.  Biol.  Chem.,  lOKi  (24).  327. 

isBer.  Deut.  Chem.  Ges.,  1914  (46)',  2327. 


6-46  DI  ABET  EH 

H  H  H  H  on  OH  Oil 

I  I  I  I  I  I  I 
IT— C— OH         C=0             0=0      H— C— OH         C=0             C=0                0=0 

II  I  I  I  I  I 
H— C— on  H— C— OH  HO— (  — H          C'=0      H— C— OH  H— C— OH  HO— C-H 

II  I  i  I  I  I 

H— C— Oil  H— C— OH  H— C— OH  H— C— OH  H— C— OH  H— C— H        IT— C— H 

I  I  I  I  I  I  I 

H  TT  II  H  H  H  H 

Glycerol  dglycerio  l-glyceric  dihydroxy         d-glyceric       d-lactic  acid.  1-lactic  acid. 

aldehyde  aldehyde  acetone  acid  ' ■: ' 

(alcohol)  (aldose)  (aldose)  (ketose)  3-carbon  saccharinic 

acid. 

Neuberg  fed  animals  and  men  considerable  doses  of  impure  d,l- 
o-lycerie  aldehyde  (glycerose)  and  saw  its  apparently  complete  util- 
ization. Parnas  demonstrated  increased  glycogen  in  tortoise  livers 
perfused  witli  d,l-glyceric  aldehyde.  Smedley  noted  the  rapid  dis- 
appearance of  glyceric  aldehyde  added  to  liver  emulsions.  Sansum 
and  AVoodyatt  fed  pure  crystalline  d,l-glyceric  aldehyde  to  rabbits 
and  guinea  pigs  in  doses  as  high  as  2.8  gm.  per  kg.  with  no  apparent 
ill  effects.  A  dose  of  5  gm.  per  kg.  in  a  rabbit  caused  diarrhea  with 
unchanged  triose  in  the  passages.  There  was  marked  diminution  of 
urine  with  albuminuria,  which  then  persisted  for  10  days.  A  dose 
of  6.8  gm.  per  kg.  killed  in  4  hours.  In  no  case  was  there  an  alimen- 
tary triosuria.  The  average  lethal  dose  hj  the  subcutaneous  route  was 
2.2  gm.  per  kg.  as  compared  with  18  gm.  of  glucose  per  kg.  in  the 
same  set  of  animals.  Suppression  of  urine  is  a  regular  manifestation, 
but  the  visceral  changes  at  autopsy  are  slight.  AVhen  d,l-glyeeric 
aldehyde  is  injected  intravenously  at  the  rate  of  only  0.15  gm.  per  kg. 
per  hour,  and  possibly  at  slower  rates,  unchanged  glyceric  aldehyde 
appears  in  the  urine,  but  no  glucose.  ( It  will  be  recalled  that  glucose 
may  be  injected  continuously  at  the  rate  of  0.8  gm.  to  0.9  gm.  per  kg. 
per  hour  without  causing  glycosuria.)  When  administered  to  diabetic 
individuals  d,l-glyceric  aldehyde  may  increase  glycosuria.  AVhen 
given  to  completely  phlorhizinized  and  glycogen-free  dogs  it  is  pos- 
sible to  demonstrate  a  quantitative  conversion  of  the  triose  into  glucose, 
the  increase  in  glycosuria  corresponding  exactly  with  the  weight  of 
glyceric  aldehyde  given.  However,  owing  to  the  toxic  effects  of  gly- 
ceric aldehyde  on  the  kidneys  there  may  be  an  incomplete  excretion 
of  all  the  sugar  formed.  The  suppression  of  urine  has  in  the  past 
been  mistaken  for  a  beneficial  effect,  since  it  may  lead  to  diminished 
excretions  of  sugar,  acetone,  aceto-acetic  and  /J-hydroxybutyric  acids. 

Einbden  and  his  coworkers  demonstrated  the  formation  of  lactic 
acid  from  glyceric  aldehyde  added  to  washed  blood  corpuscles.  The 
keto-triose,  dihydroxyacetone,  was  observed  to  produce  less  lactic  acid, 
but  otherwise  it  is  not  improbable  that  the  behavior  of  the  ketotriose 
is  analogous  to  that  of  the  aldo  foi-ms.  Thus  AFostowski  found  dihy- 
droxyacetone to  be  a  glycogen  forinei-.  and  Kingei- "''  reported  its  com- 

iiKingcr  and  Fraiikol,  Jour.  V,\o\.  (  licm.,   \\n\    (18),  233. 


THE  sTATi:  or  Tin:  si  <;.\i;  i\  riii:  lu.oon  G47 

plt'tc  1 1'aiist'oi'iiiat  ion  into  glucose  in  tlic  i'ull\-  plilorliizini/.cd  (1(»<j:. 
The  complete  conversion  of  d,l-glyceric  aldehyde  into  glucose  in 
phlorhizinized  dogs — its  transformation  into  glycogen  in  the  perfused 
liver,  its  disappearance  as  such  when  added  to  liver  emulsions,  all 
indicate  that  glyceric  aldehyde  (like  diose  and  other  sugars  in  gen- 
eral) is  converted  into  glucose  in  the  body  as  a  preliminary  step  in 
utilization.  The  fact  that  large  doses  may  be  given  by  the  alimentary 
route  without  causing  melituria  or  death,  whereas  much  smaller  doses 
given  subcutaneously  may  prove  lethal,  together  with  the  very  low 
rate  at  which  glyceric  aldehyde  has  to  be  given  by  vein  in  order  not  to 
produce  melituria,  all  point  to  the  liver  (and  bowel  wall)  as  the  chief 
sites  of  its  conversion.  Glj-ceric  aldehyde  has  figured  prominently  in 
theories  of  the  normal  catabolism  of  glucose,  and  on  the  basis  of  his 
observations  concerning  the  formation  of  lactic  acid  from  this  triose 
by  blood  corpuscles  Embden  regards  it  as  a  chief  normal  intermediate 
substance  in  the  oxidation  of  glucose  in  the  cells.  Now  glucose  may 
be  oxidized  in  the  body  at  the  rate  of  0.6  gm.  per  kg.  per  hour  under 
suitable  circumstances,  and  if  eveiy  molecule  of  glucose  oxidized  were 
first  split  to  give  two  molecules  of  glyceric  aldehyde,  as  the  Embden 
hypothesis  would  demand,  then  glyceric  aldehyde  would  be  formed 
in  the  body  at  the  rate  of  0.6  gm.  per  kg.  per  hour,  and  the  place  of 
formation  would  be  within  the  cells  of  the  body  at  large,  the  muscles 
representing  the  most  important  sites  of  oxidation.  However,  if 
glyceric  aldehyde  is  introduced  into  the  systemic  blood  at  only  one- 
fourth  of  this  rate,  unchanged  triose  appears  in  the  urine  and  may  be 
demonstrated  in  the  blood.  But  glyceric  aldehyde  has  never  been 
found  in  the  blood,  urine  or  tissues  under  any  other  circumstances. 
Olyceric  aldehyde  may  of  course  enter  the  body  via  the  portal  route 
at  faster  rates  without  causing  triosuria,  but  then,  as  stated,  it  would 
appear  not  to  be  oxidized  directly  but  first  assimilated,  i.e.,  trans- 
formed into  glucose.  Recently,  for  other  reasons,  von  Flirth  ^"  has 
also  questioned  the  tenability  of  Embden 's  hypothesis. 

Lactic  acid  from  triose:  \\'lien  alkali  acts  on  (rlncose  (or  licxoscs  in  srcTieraH 
in  the  absence  of  oxycren,  lactic  acid  is  formed  in  amounts  as  hipli  as  40  to  60 
per  cent,  of  the  weifrlit  of  the  sugar  used,  provided  the  conditions  are  propcrlv 
controlled.  Hut  in  tlie  y)resence  of  sufficient  oxyiren  no  lactic  acid  is  formed. 
Still,  ]n-cformed  lactic  acid  will  not  be  destroyed  if  it  is  added  to  this  latter 
mixture.  So  it  is  clear  that  lactic  acid  is  not  an  intermediate  in  the  oxidative 
breakdown  of  su<iars  in  the  alkaline  solution.  Meisenlicinicr  accord<nurly  sutr- 
gested  the  obvious  prol)ability  that  there  was  some  labile  i)rccursor  i>f  lactic 
acid  which  burned  in  the  presence  of  oxy<ren :  in  the  absence  of  oxyfren,  rearranged 
to  give  lactic  acid.  He  proposed  glyceric  aldehyde  as  such  a  body.  Xef.  how- 
ever, holds  that  the  immediate  precursor  of  lactic  acid  is  methyl  glyoxal 
(CH3  —  CO  —  COH).  which  forms  lactic  acid  bv  undergoing  what  is  known  to 
chemists  as  a  "henzilic  acid  rearranuenient."'  These  phenomena  are  remarkably 
similar  to  those  that  occur  in  tlie  body. 

One   other   important   point    should  be   emphasized   in   this   place.     The   trioses 
condense  in  the  i)resence  of  alkali   to  yield  among  other  things  certain   hexoses. 

iTBiochem.  Zeit..  1916    (69),   199. 


648  DIABETES 

ami,  as  dest'rilicd  under  hcxoscs — any  sujiar  of  tliat  type  will  in  tlu'  ))ioseiire  of 
alkali  enter  into  a  complex  etjuilibriiim  with  several  other  hexoses.  Any  of  these 
may  again  split  into  3-carl)on  compounds  such  as  the  trioses  and  then  again  con- 
dense, and  so  on,  as  long  as  they  do  not  become  converted  into  lactic  acid 
or  the  saccharinic  acids — substances  which  are  not  reconvertible  into  sugar. 
Nef  formulated  the  view  that — were  it  not  for  the  occurrence  of  these  irre- 
versible reactions — any  sugar  in  the  presence  of  alkali  would  come  finally  to 
represent  an  equilibrium  of  every  possible  sugar  of  2  to  6  carbon  atoms  {i.  e.,  56) 
together  with  all  of  the  myriad  intermediate  forms.  In  the  body,  however,  lactic 
acid  can,  be  converted  into  sugar.  So  this  l)ar  to  the  great  equilil)rium  is  there 
nonexistent,  and  it  is  conceivable  that  in  tlie  body  there  actually  exists  an  c()uili- 
britun  of  this  sort. 

Ill  all  of  Embden's  experiments  there  was  a  lack  of  oxygen,  so 
that  the  phenomena  //(  vii'o  and  in  alkaline  solution  in  vitro  are  strik- 
ingly parallel.  Yet  Embden,  on  the  basis  of  these  experiments,  re- 
gards sareolactic  acid  {i.  e.  d-lactic)  as  a  chief  normal  breakdown 
product  of  glucose  in  the  body  over  the  glyceric  aldehyde  route.  But 
it  is  hard  to  see  why  this  assumption  is  any  more  rational  than  it 
would  be  to  say  that  lactic  acid  is  an  intermediate  in  the  oxidative 
breakdown  of  sugars  in  the  alkaline  solution  outside  the  bod}',  which 
it  certainly  is  not.  Although  lactic  acid  will  disappear  from  a  sur- 
viving asphyxiated  muscle  if  ox^^gen  be  resupplied  to  it  (Fletcher)  ^^ 
and  although  this  disappearance  will  not  occur  in  an  alkaline  solution 
experiment,  unless  some  stronger  oxidizing  agent  than  ILO2  is  used, 
still  it  must  be  remembered  that  the  asphyxiated,  lactic  acid-contain- 
ing muscle,  has  acquired  an  acid  reaction.  If  the  alkaline  solution  be 
acidified  and  a  trace  of  ferrous  sulphate  added  it  also  permits  lactic 
acid  to  burn  with  peroxide,  and  still  we  know  that  the  lactic  acid  was 
not  an  intermediate  until  the  oxygen  supply  became  deficient.  Lactic 
acid  is  probably  an  intermediate  in  the  sugar  catabolism  only  during 
relative  asphyxia. 

All  the  substances  whose  formula?  are  given  above  have  been  shown 
to  be  capable  of  conversion  into  glucose  in  the  body.  The  details 
of  the  stej)s  involved  have  not  been  established,  but,  in  conformity 
with  the  chemical  theories  developed  by  Nef  and  discussed  under 
liexoses,'  the  transformation  of  these  substances  into  glucose,  as 
well  as  their  occurrence  in  the  course  of  its  breakdown,  are  best  ex- 
])lained  on  the  basis  that  all  of  them  participate  in  the  same  great 
chemical  equilibrium  with  the  sugars,  and  that  this  participation  de- 
])eiids  upon  their  dissociation  into  unsaturated  residues.  These  resi- 
dues* are  in  dynamic  chemical  eciuilibrium  with  the  molecules  from 
wiiicli  they  are  derived  and  with  those  derived  from  sugars.  When 
there  is  a  rapid  loss  of  glucose  from  the  body  these  substances  tend  to 
become  glucose,  in  accordance  willi  llie  laws  of  chemical  equilibrium. 

TETROSES 

Tliere  are  six  possible  tetroses  (4  aldo-  and  2  keto-").  The  entire 
subject  of  their  physiology,  which  has  undoubtedly  considerable  bio- 

17  Jour,  of  Physiol.,  1907    (35),  247. 


PEXTOSHS  649 

logic  importance,  is  virtually  a  clean  page.  Tlicy  have  never  l)een 
found  iu  the  urine,  since  there  are  at  present  no  established  methods 
for  their  detection,  and  ett'orts  have  been  lacking. 

PENTOSES 

Chemical  theory  demands  the  existence  of  fourteen  pentoses,  i.  e., 
six  aldo-pentoses,  four  2-keto-pentoses  and  four  3-keto-pentoses.  Only 
those  better  known  to  cliemists  have  received  biological  study,  e.  g., 
arabinose  and  xylose.'"*  Of  these  the  optically  inactive  or  d-  1-arabin- 
ose,  tlie  1-arabinose  and  1-xylose  are  the  best  known.  When  even 
small  quantities  of  pentose  gain  access  to  the  circulating  blood,  pentose 
is  excreted  in  the  urine.  Ebstein  ^'■'  reports  the  appearance  of  traces 
in  the  urine  of  a  man  (in  which  none  had  been  previously  demon- 
strated), after  the  administration  of  so  little  as  0.25  gram  of  1-ara- 
binose b}'  mouth.  Bergell  -"  found  reactions  for  pentose  in  the  urine 
seven  to  ten  minutes  after  ingestion  of  the  same  sugar,  and  when  given 
subcutaneously,  Fr.  Voit  -^  saw  about  50  per  cent,  excreted.  Neu- 
berg  and  "Wohlgemuth  --  gave  a  normal  man  15  grams  of  d-  1-ara- 
binose and  recovered  4.5  grams  of  d-,  and  only  1.04  grams  of  1-ara- 
binose in  the  urine.  On  the  other  hand  1-arabinose  becomes  converted 
in  part  into  the  dextro-form,  since  both  forms  appear  in  the  urine 
when  only  one  is  given.  Xylose  has  been  found  to  behave  in  general 
like  arabinose.-^ 

Since  all  writers  agree  that  10  to  50  per  cent,  of  administered  pen- 
toses are  excreted  in  the  urine  even  when  given  per  os  in  very  small 
quantities,  and  since  pentoses  occur  in  many  foods  (plums,  cherries, 
apples,  etc.)  or  result  from  the  bacterial  decomposition  of  other  carbo- 
hydrates, it  is  inevitable  that  alimentary  pentosurias  should  occasion- 
ally occur  in  nearly  everj'  normal  individual,  and,  as  a  matter  of  fact, 
most  normal  urines  give  reactions  wdiicli  indicate  the  presence  of 
some  pentose  { Cremer,  Funaro,  Cominotti).  Vice  versa,  one  may 
conclude  that  very  little  pentose  normally  occurs  in  the  blood,  since 
otherw'ise  more  of  it  w'ould  appear  in  the  normal  urine  than  does ;  and 
finally,  that  pentoses  must  play  but  a  minor  role  in  the  general  metabo- 
lism of  the  carbohydrates.  Therefore  it  is  highly  improbable  tiiat 
during  the  breakdown  or  synthesis  of  glucose  in  the  body  the  hexoses 
split  to  any  extent  into  a  pentose  and  formaldehyde.  The  same  holds 
good  for  the  behavior  of  hexoses  in  the  presence  of  alkali.  They 
split  almost  exclusively  into  chains  of  2,  3  and  4  carbon  atoms  (Nef). 

18  Rhamnose  is  a  methyl  pentose,  representing  a  elass  of  substances  closely 
related  to  the  pentoses. 

i9Vircho\v"s  Archiv.,  18n2    (120),  401. 

2oFestschr.  f.  E.  v.  Levden,  1<)02   (2),  401. 

2iDeut.  Arch.  f.  klin.'Med.,  ISO?    (58),  ,523. 

22Zeit.  f.  physiol.  C'hem.,  1002   (35),  41. 

23  For  literature,  see  Xeuberg.  "Der  Harn  sowie  die  iibrigen  Ausscheidiingen, 
etc."      (Springer,  Berlin,  1911),  I,  p.  370. 


650  diabhtes 

chronic  pentosuria  24 
Tilt'  literatuiv  coutaiiis  repoi'ts  of  some  30  cases  in  which  consid- 
erable (jnantities  of  pentose  have  Won  excreted  steadil}^  in  the  urine 
regardless  of  the  character  or  (inantity  of  the  food.  Even  during  a 
fast  the  quantity  excreted  has  remained  virtually  constant  in  some 
cases.  Outputs  as  high  as  36  grams  per  day  have  been  recorded. 
Such  quantities  of  pentose  could  not  be  introduced  into  the  body  from 
without  by  any  known  means  without  causing  pentosuria  of  marked 
degree.  Accordingly,  in  some  cases  there  is  either  an  overproduction 
of  endogenous  pentose,  or  an  abnormal  entry  into  the  blood  stream 
of  pentose  which  is  normally  bound  in  the  tissues.  The  process  would 
then  be  analogous  to  that  in  which  lactose  from  the  mammary  glands 
occasionally  gains  access  to  the  general  circulation  and  appears  in  the 
urine.  This  conclusion  is  confirmed  by  the  work  of  Bial,  Blumenthal 
and  Tintemann,  who  found  that  certain  pentosurics  displayed  no  les- 
sened tolerance  for  administered  pentose.  The  origin  of  the  pentose 
is  unknown.  Nucleo-protein  of  cell  nuclei,  and  galactose  have  been 
suggested  as  possible  sources.  The  disease  has  been  found  in  differ- 
ent members  of  the  same  family  and  appears  to  be  a  harmless  anomaly. 
The  pentose  found  in  the  urine  in  cases  of  all  types  has  sometimes 
been  reported  as  optically  inactive,  sometimes  as  dextro-  or  levo- 
rotatory:  1-arabinose  (dextro-rotatory),  d-xylose  and  d-xyloketose 
appear  to  have  been  identified.-"' 

HEXOSES 

Chemical  Introduction. — Structural  theory  demands  the  existence  of  32 
isoiiicric  liexose  su<>ars  of  the  formula  CoH,„0„.  The  behavior  of  the  hexoses 
when  dissolved  in  very  dilute  alkali  makes  it  convenient  to  consider  tliem  in  four 
natural  series  of  oifrht  members  each.  Thus  one  series  comprises  tlic  S  hexoses 
whose  structural  formuUie  ai)pear  below.     This  may  be  called  tlie  d-filucose  series. 

(1)  (2)  (3)  (4)  (5)  (6)  (7)  (8) 

CHO  CHO  CHiOH        CHO  CHO  CH.OH        CH^OH  CHoOH 

H-i-OH  HO-C-H               CO  H-C-OH  HO-C-H  CO  H-COH      HO-C-H 

I                      I                        I                     I                      I  I                     I                         I 

HO-C-H      HO-C-H      HO-C-H  H-C-OH      H-C-OH  H-C-OH          CO                   CO 

I  I                    I                  I                   I  I                  !                     I 
H-6-0H      H-C-OH      H-C-OH  H-C-OH      H-C-OH  H-C-OH  H-C-OH       H-C-OH 

II  I  I  I  I  I  I 
n-(^'-OH      H-r-OH      H-COH       H-C-OH      H-C-OH    H-C-OH    H-C-OH       H-C-OH 

r  r         1  I  I  I  I 

CH,OH         Cn.OH         CH.OH        CH,OH         Cn,OH        CH.OH        CH.OH  CHsOH 

d-i)seudo 
dglucase     d-mannose     d-fructose     d-allose  d-latose       fructose     ad-ghitose        /3-d  ghitose 

Tliere  is  also  an  l-<:lucose  series  in  which  the  members  are  the  mirror  imajres 
of  the  above.  There  is  a  tliird  series  coniprisin<r  d-galactose,  d-talose,  d-tajiatose, 
l-sorl)ose,  1-idose,  l-<;ulose  and  alplia  and  beta  d-galtose;  and  a  fourtii  series  whose 
relatioiisliip  to  the  d-<ralaclose  series  is  tlie  same  as  that  of  the  l-jrhicose  to  the 
d-glucose  series.     Consideration   of  tlie  d-<ilucose  series  will   liriiijj:  out    tlie  priii- 

•.;4  See  fiarrod.  "Inliorn  I'Jrors  of  Mctaliolism."  (~)xfor(l  :Mcd.  Pulil.,  I'.IO'.l;  l.iuicct. 
.lulv,  1 !)()!». 

■^■^Vor  literature  see  lliller,  Jour.  Biol.  C'liem.,   VMl    (30),  120. 


iij:\<>s/:s  G,')! 

ciplos  coininoii  to  all.  l-Aiun'malion  of  tlic  S  toiiimhu'  sIkiws  tlial  iiiiinhLTs  1  2, 
4  and  .>  have  aldfliydc  fjroiijjs  (  11 — C^ O)  at  tlic  end  of  the  eliaiii  and  arc  licncc 
aldolii'xoscs.  Mcndici's  :?  and  t>  have  kctom-  (C^f))  >rroii,))s,  at  tlic  second  car- 
bon atom,  and  are  therefore  2-keto-liexoses,  while  numbers  7  and  S  havinjj  ketone 
jjroups  at  the  third  carbon  atom  are  .'5-keto-hcxoses.  Since  each  series  of  S  sugars 
has  a  like  nnmber  of  the  diircrcnt  types  there  are  in  all  Iti  aldoln-xoscs.  ci^lit  2-keto- 
and  ciiiht  ;i-keto-liexoses. 

Jt  had  lonjr  been  known  that  if  a  solution  of  any  optically  active  su^'ar.  such 
as  d-jihicose.  was  alkalini/.ed  the  solution  <iradually  lost  its  optical  activity.  It 
was  later  shown  by  Lol)rv  de  Bruyn  and  van  Ekcnstein  that  a  solution  of 
d-jrhu'ose  in  very  dilute  alkali  coines  to  contain  a  tjroup  of  4  hexoses  in  dynamic 
chemical  equilibiiuni.-'i-  Ncf  -"  held  that  in  such  solutions  there  is  an  e(piili- 
brium  of  at  least  eiijlit  hexoses  as  above  depicted.  Any  one  of  these  sugars  when 
placed  in  alkali  reproduces  the  other  seven,  since  the  members  of  the  series  are 
reciprocally  convertible  one  into  another.  The  same  holds  good  for  the  members 
of  the  d-galactose  series  and  for  tlu^  1-galactose  and  l-glucoae  series.  Rut  the 
reciprocal  transformation  of  tlie  members  of  one  series,  such  as  d-glucose,  into  a 
hexose  of  another  series,  such  as  d-galactose.  occurs,  if  at  all,  only  to  a  minute 
degree,  because  sucli  transformation  involves  the  breaking  of  the  hexose  chain  into 
2,  '.i  and  4  carbon  fraiiinents  with  subse(|uent  recombinations,  and  wlien  this 
occurs  irreversi])le  reactions  are  prone  to  intervene.  The  formation  of  lactic 
acid  and  the  saccharines  aie  rej)resentative  of  these  irreversible  reactions.  (In 
the  body,  however,  glucose  may  be  converted  into  lactic  acid  in  the  muscles  and 
elsewhere,  whereas  in  diabetes  lactic  acid  can  be  converted  easily  back  intc 
glucose;  in  diabetes  galactose  is  convertible  into  glucose,  etc.,  so  that  in  the 
body  the  transformation  of  hexoses  of  ditferent  groups  one  into  another  offers  no 
difficulty. ) 

In  order  to  explain  the  effects  of  dilute  alkali  on  hexoses  just  described,  some 
conception  of  labile  intermediate  products  is  a  logical  necessity,  for  when  levulose 
changes  into  glucose  there  is  necessarily  some  intermediate  phase.  The  nature  of 
these  phases  has  been  the  subject  of  study  by  many  chemists,  and  this  study 
involves  always  the  question  of  sugar  dissociation. 

Sugars  are  weak  acids.  They  form  salts  with  metals,  and  Cohen  28  and  later 
Michaelis  and  Rona  20  have  determined  by  physico-chemical  methods  the  ioniza- 
tion constants  for  glucose  and  other  sugars.  Sugars  are  also  polyatomic  alcohols, 
and  either  aldehydes  or  ketones.  Sugar  chemistry  reverts  to  the  chemistry  of 
these  three  classes  of  compounds. 

A.  P.  Mathews  so  and  ]Michaelis  have  suggested  that  the  effect  of  alkali  on  a 
siiffar  such  as  srlucose  is  to  increase  enormouslv  the  concentration  of  the  gh;cose 
anion,  i.  e.,  KOH  leads  to  the  formation  of  K-glucosate  (see  Fig.  3,  p.  21),  which, 
being  the  combination  of  a  powerful  base  with  a  very  weak  acid,  has  a  high 
electrolytic  dissociation  constant.  These  anions  accordinir  to  this  view  are  sub- 
ject to  cleavages  and  intramolecular  rearrangements.  Nef  also  holds  that  the 
first  effect  of  the  alkali  (c.  jr.,  KOTI)  is  to  form  a  salt,  but  his  far  more  detailed 
conception  of  the  subseciuent  changes  which  lead  to  reciprocal  transformations 
of  hexose  sugars  one  into  another,  involves  other  principles  which  represent  the 
outsiTowth  of  his  earlier  work  on  the  properties  of  simpler  aldehydes  aiul  ketones. 

These  reciprocal  transformations  are  dependent,  according  to  Xef,  ujion  the 
aldehydic  or  ketonic  character  of  the  sugar,  whereas  the  oxidative  phenomena  and 
the  saccharinic  acid  formation  to  be  described  presently,  depend  upon  the  alcohol 
groups.  The  principles  involved  can  best  be  understood  if  we  first  consider  the 
behavior  of  a  simple  aldehyde  (acetaldehyde)  and  a  simple  ketone  (acetone). 
Acetaldehyde  in  the  presence  of  water  forms  a  hydrate  (comparable  to  chloral 
hydrate) . 

"oRec.  trav.  cliim.  de  Pays  Pas  (14),  l-'iS  and  203:  d.",),  92:  (IG).  2r,7:  (10). 
1  and  10. 

2- Liebig's  Annalen.   1007    (3o7).  204:    1010    (37G),   1. 
2sZeit.  f.  phvsikal.  Chem.,  1001    (36),  60. 
20  Biochem.  Zeit.,  1012   (47),  447. 
30  Jour.  Biol.  Chem.,  1909  (6),  1. 


652  DIABETES 

H  H 

CH3— c  =  0  +  n  on  ^  ciia— c— OH 


ok 


This  hydrate  possesses  ioiiizable  hydrogen  in  its  OH  groups,  and  in  the  presence 
of  a  metallic  liydroxide,  MOH,   can  accordingly  form  a  salt    (comparable  to  an 

H 

alcoholate)  ;   tiius:   CHj — C— OH, 

and  this  salt  being  highly  dissociable  falls  apart  into  MOH  and  a  "methylene 
enoV  (in  this  case  hj'droxyethylidene)  : 

H 

1  *  I 

CH3— C— OH  ^  CH3— C— OH  +  MOH. 

I  1 

OM 

(Herein  lies  the  point  of  departure  of  Nef's  view  from  the  foregoing.)  This 
methylene  enol  then  rearranges  to  form  the  "olefine  dienol"  CHo  =  CHOH  (in  this 
case  vinyl  alcohol).  Ketones,  on  the  other  hand,  form  the  oletine  dienol  directly 
without  forming  the  methylene  enol.  In  the  case  of  KOH  and  acetone  (dimethyl 
ketone)  there  is  the  same  formation  of  the  hydrate  followed  by  salt  formation  and 
the  loss  of  KOH,  but  the  latter  does  not  all  come  from  one  C  atom  as  it  does 
when  split  out  of  aldehydes,  thus: 

OH  OK 

1  I  II 

CH3— C— CH3  +  KOH  ^  CH3— C— CH3  +  ILO  <^  GH3— C— CH2 

OH  OH  OH 

( hydrate  of  acetone ) 

In  a  manner  entirely  analogous  to  what  occurs  in  the  simple  aldehydes  and 
ketones,  the  two  aldohexoses,  glucose  and  mannose,  and  the  ketohaxose  levuloso, 
can  form  one  and  tlie  same  enol  molecule.  And  vice  versa  this  enol  molecnli  may 
open  its  double  bond  in  two  ways  as  shoAvn  below  (  (a),  (b)  and  (c)  )  and  the 
dissociated  molecules. 


(a) 

OH 

1 

1 

(c) 

OH 

1 

I 

(b) 
OH 

(fl) 

OH 

(e) 

H 

1 

0- 

I 

-C— 

1 

1 

H- 

1 
-0 

1 1 

H- 

-C- 

1 

- 

H- 

-0 

H- 

I 
-C- 

-OH 

HO- 

I 

-f— 

1 

- 

2 

1 1 

r- 
1 

-OH 

1 
-C- 

1 

-OH 

C— OH 

1 

r- 

1 1 

-OH 

HO- 

1 

-c- 

1 

-H 

.3 

HO- 

1 

-c- 

t 

-II 

HO- 

I 

— c- 

I 

-H 

H- 

1 

— C- 

1 

-OH 

H 

II 
r- 

1 

-OH 

H- 

1 
-c- 

1 

-OH 

4 

II- 

1 
1 

-OH 

II- 

1 
— ('- 

-OH 

II- 

-('— OH 
1 

H- 

1 

-r- 

1 

-on 

H- 

1 

-r- 

1 

-OH 

5 

H- 

1 
-r- 

-on 

H- 

-c- 

1 

-on 

H- 

I 
1 

-on 

H- 

1 
-r- 

-on 

H- 

1 
-c- 

-OH 

G 

H- 

-r- 
1 

-on 

H- 

1 
— r- 

-OH 

II- 

1 

— r- 

1 

-on 

H- 

1 

-OH 

H 

1 

H 

( 

a  1.  2 

li 

Enol  niolcculi' 
d-glucosi"  oleiiiio 

dienol) 

I 

H 

1 

H 

HEX08ES 


65:^ 


(a)  and  (b)  will  be  in  dynamic  equilibrium  with  the  enol  (c).  Now  if  II  and 
OH  are  again  taken  on  by  (a)  and  (b)  this  assumption  of  the  elements  of  water 
can  take  place  in  thrt-c  different  ways,  to  regenerate  d  Icvulose,  d-nianiiose  and 
4-glucose.     Thus  if  in  the  case  of   (a),  Oil  is  added  to  carbon  atom  number  one 

OH 

I 
tins  will  form  tlie  group  H — C — OH  whicli  represents  a  hvdrated  aldeiivde  "roup 

I  .  .        »       1 

and  will  lose  water  to  become  CHO.  'ihen  H  going  to  the  second  C  atom 
completes  the  formula  of  d-mannose.  In  a  similar  way  (b)  can  form  d-glucose. 
But  if  tlie  OH  went  to  the  second  carbon  atom  this  group  would  tliereby 
bi'come   a   hydrated   ketone   similar   to   the   hydrate   of   acetone   and    lose    water 

to  form  0  =  0,  while  H  going  to  the  end  carbon  atom  woukl  coiiiplete  tiie  formula 

of  d-Icvulose.  In  a  manner  entirely  analogous  there  is  an  enol  molecule  which  is 
common  to  d-allose,  d-latose  and  d-pseudo  fructose  (see  (d)  above). 

It  will  be  noticed  that  each  of  these  enols  is  in  equilibrium  witli  a  2-keto- 
hexose  and  two  aldo-hexoses.  Now  these  2-keto-hexoses,  d-fructose  and  d-pseudo 
fructose,  in  accordance  with  general  ketone  behavior,  are  capable  of  yielding  an- 
other common  enol,  /.  e.,  a  2-3  enol  (with  the  double  bond  between  tlie  second  and 
third  carbon  atoms)  as  represented  at  (e)  and  this  2-3  enol  (by  a  process  like 
that  just  detailed  for  the  1-2  enols)  can'  accoimt  for  the  formation  of  the  two 
3-keto-hexoses  whose  formulae  are  given  above.  The  same  general  principles  hold 
for  each  of  the  series  of  hexoses  ( For  further  elaboration  of  the  theory 
see  Nef's  original  papers.)  A  similar  use  of  enol  molecules  in  this  connection 
is  made  In'  Neuberg. 

It  remains  now  to  point  out  that  if  to  a  simple  aqueous  solution  of  sugar, 
oxygen  be  supplied  in  the  form  of  air  or  HoO^,  no  oxidation  occurs.  But  if  the 
solution  be  alkalinized  then  the  sugar  is  readily  burned.  In  the  absence  of 
oxygen  and  the  presence  of  alkali  somewhat  stronger  than  that  foimd  most 
favorable  for  the  reciprocal  transformation  above  detailed,  there  occur  certain 
irreversible  reactions  such  as  the  formation  of  lactic  acid  and  the  so-called 
saccharines.  When  an  alkaline  sugar  solution  is  treated  with  oxygen  it  yields 
COo,  H„0,  and  formic,  glycollic,  glyceric  and  certain  trihydroxy-butyric  and 
hexonic  acids,  depending  on  the  sugar  used.  Witliout  oxygen,  or  witii  too  little 
oxygen,  lactic  acid  and  the  saccharinic  acids  make  their  appearance  (cf.  the  forma- 
tion of  lactic  acid).  The  explanation  of  these  phenomena  rests  in  tlie  conception 
that  alkali  increases  the  dissociation  of  sugar,  and  tliat  tlie  dissociated  fragments 
burn  or  rearrange  depending  upon  tlie  conditions  of  the  experiment. 

In  this  connection,  according  to  Nef,  we  are  dealing  with  the  alcohol  groups 
of  the  sugars  and  may  advantageously  turn  for  a  moment  to  the  properties  of 
inethvl  alcohol.     This  substance  consists  imder  ordinarv  circumstances  of  a  great 


Glucose 
(1) 

O 

II 
C— H 

I 
H— C— OH 

I 
OH— C— IT 

I 
H— C— OH 

I 
H— C— OH 

H— C— OH 

H 


Glucose  ion 

(2) 

(— ) 

0 

II 

C— H 

I 
H— C— 0— 

I 
OH— C— H 

I 
H— C— OH 

H— C— OH 

I 
II— C— OH 

I 
H 


(  +  ) 
— H 


K-glucosate 
(3) 

O 

II 
C— H 

I 
H— r— OK 

I 

OH— C— IT 
I 
H— C— OH 

I 
H— C— OH 

I 

H— C— OH 

I 
H 


Metliylene 
particle. 
(4) 

O 

II 
C— H 

I 
— C— 

I 
no— C— II  +  KOH 

r 

H— C— OH 

I 
H— C— OH 

II— C— OH 

I 
H 


654  D I. \  BETES 

]>i('lMHi(lcianco  <if  iiiHlissociali'd  nuili'ciiles  in  dynamic  ('(|uilil)riiim.  witli  a  very 
minute  quantity  of  dissociated  methylene  CIIiOH  ^  C'lL  +  ILO.  Tlie  primary 
efi'ect  of  alkali  (KOH)  is  to  form  a  salt,  ('H3 — 0\\  (or  K-metliy!ate) ,  wliioli  being 
liiiilily  dissociable  breaks  down  to  give  CH^  and  KOH.     Tlie  proportion  of  free 

mi'tliylene  is  thereby  enormously  increased,  ^^'hat  tlien  befalls  the  metiiylene 
will  depend  on  the  amount  of  oxygen  present  and  on  the  various  other  factors 
which  enter  into  the  conditions  of  tlie  experiment.  These  general  principles  are 
applicable  directly  to  the  j)olyatomic  alcohols — the  hexoses  and  other  sugar.s — as 
shown  on  preceding  j)age. 

In  the  presence  of  sullicient  oxygen  the  methylene  particle  takes  on  oxygen 
to  form  first  an  osone.  In  the  absence  of  oxygen  it  vmdergoes  intramolecular 
rearrangements,  the  details  of  which  need  not  here  be  entered  into.  It  is  these 
wliich  gives  rise  to  the  6  carbon  acids  known  as  the  saccharines  or  saccharinic 
acids. 

GALACTOSE 

A  normal  individual  weighino-  75  kilos  may  eat  about  50  grams  of 
galactose  and  show  but  a  trace  of  melituria.  ]\Iore  than  this  is  likely 
to  cause  the  presence  of  measurable  amotnits  of  galactose  in  tlie 
urine,  the  alimentary  tolerance  limit  for  this  sugar  being  therefore 
about  0.6  to  0.8  grams  per  kilogram  of  body  weight.  AVe  have  no 
direct  data  concerning  the  time  within  which  50  grams  of  galactose 
are  absorbed  by  a  man  of  average  weight.  When  given  intravenously 
at  uniform  rates,  unchanged  galactose  appears  in  the  urine  of  dogs 
receiving  slightly  more  than  0.1  gm.  per  kilo  per  hour.  The  tolerance 
for  galactose  appears  to  be  lessened  in  phos])h()rus  poisoning  and  in 
many  other  conditions  which  cause  apparent  parenchymatous  changes 
in  the  liver,  so  that  after  administration  of  50  grams  of  galactose  by 
mouth,  as  much  as  10  to  12  grams  may  be  excreted  in  the  urine. 
(Bauer.)  On  the  other  hand,  ligation  of  the  common  duct  does  not 
lessen  the  tolerance  for  galactose  in  rabbits  (Reiss  and  Jehn,  Hierose) 
so  that  the  lowered  tolerance  following  phosj^horus  administration  ap- 
l)eai's  to  be  indepentlent  of  the  disturbed  biliary  function.  Infants 
suffering  from  gastro-enteritis  may  show"  alimentary  lactosuria,  and 
along  with  the  lactose  some  of  its  constituent  galactose  may  apear  in 
the  urine.  The  question  thus  naturally  arises  as  to  whether  the  lowered 
tolerance  for  galactose  in  phos])horus  poisoning  and  other  liver  dis- 
eases may  not  be  due  to  an  increased  permeability  of  the  intestinal  wall, 
or  to  changes  elsewhere  in  the  body  besides  the  liver.  Worner  found 
that  galactose  injected  directly  into  the  portal  vein  Avas  handled  by 
liealthy  and  phosphorized  rabbits  in  the  same  relative  prop(n-tions  as 
when  giv(Mi  to  these  animals  by  mouth,  tlnis  ai)])ai'eii1ly  (>xcluding  the 
bowel  as  a  (M)iiti-il)iit('f  to  the  d(M're;ised  tolerance.  It  is  unlikely  also 
lliat  1li('  ki(hieys  in  pliosplioi'izcd  animals  wei'c  iHMulci'ed  abnoniially 
))t'nneal)h'  for  galactose,  since  when  the  kidneys  alone  are  ))liosi)liorized 
without  affeeting  the  liver,  the  excretion  of  galactose  after  adminis- 
tration l)y  mouth  oi-  into  a  xcin  lias  been  retarded  rather  than  ha.stened. 
These  princii)les  ha\c  iK'couie  ineorjioi-ated  in  a  clinical  test  for  dis- 
ease of  the  lie|»;itie  | )a I'eneliv ma.     When  galactose  is  administered  to  a 


/. /;  17  /.0,S7v    (  Fh'CCTOSE )  655 

fully  (liahctic  <iiiiiii;il  it  is  c-apiihlc  (if  l)('iii<i:  coiivcrlcd  (iiiiiiililat  i\cl\' 
into  {i'lucosc.  h].\istiiijr  data  imlicalc  that  j^alaetoso,  like  diusi-  and 
ji'lycoiMC  aldi^liydc,  is  ciiiefly  convcrtt'd  into  itlufosc  bcfoi'c  fiirllirr  nlil- 
ization,  and  that  this  pi'occss  is  carried  out  niaiidy  in  tlic  li\cr  and 
howcl  wall.     The  litci'atni'c  of  the  sulijcci  is  <:iv('n  hchiw.'" 

LEVULOSE    (FRUCTOSE) 

The  uroup  of  ci^ht  sugars  formed  by  levulose  in  the  presence  of 
alkali  includes  glucose,  and  any  member  of  this  group  will  produce 
all  the  others.  Then,  in  cases  of  glycosuria  with  alkaline  urine 
(whether  physiological  or  due  to  medication  or  to  bacterial  decompo- 
sition), levulose  might  be  expected  to  occur  along  with  glucose.  May 
and  Koenigsf eld  have  reported  instances  of  this  ' '  urinogenous  levu- 
losuria. "  However,  since  the  OH  concentration  of  fresh  urine  seldom 
or  never  attains  a  height  sufficient  to  turn  phenolphthalein  red,  Mag- 
nus-Levy doubts  the  correctness  of  these  observations. 

Alimentary  Levulosuria. — The  tolerance  of  a  normal  body  for  levu- 
lose given  per  os  is  variable.  Doses  of  50  to  70  gm.  in  man  cause  a.s  a 
rule  no  levulosuria,  but  more  is  likely  to  do  so.  Animals  in  wiiich 
the  liver  parenchyma  has  been  damaged  by  phosphorus  are  said  to 
have  a  lower  tolerance.  In  many  other  diseases  of  the  liver  the  same 
holds  true,  and  IT.  Strauss  believed  this  fact  could  be  made  the  basis 
of  a  clinical  test  for  liver  function.  Naunyn,  however,  emphasi;^es 
the  fact  that  in  certain  cases  of  cirrhosis  of  the  liver  with  collateral 
anastomoses  between  the  portal  vein  and  vena  cava,  there  is  also  a 
lessened  tolerance  for  levulose  given  by  mouth,  owing  to  the  fact  that 
levulose  then  enters  the  general  circulation  without  having  entered 
the  liver.  As  far  back  as  1871  Eichhorst  showed  that  levulose  intro- 
duced per  rectum,  i.  e.,  where  it  will  presumably  enter  an  hemor- 
rhoidal vein  after  resorption,  is  more  likely  to  cause  alimentary 
levulosuria  than  when  swallowed,  because  of  the  extra-hepatic  an- 
astomoses between  the  hemorrhoidal  veins  and  the  vena  cava.  As  in 
the  case  of  diose,  glyceric  aldehyde  and  galactose,  intravenous  injection 
of  levulose  produces  levulosuria  in  dogs  when  tlie  rate  of  injection  is 
between  0.1  and  0.2  gram  per  kilo  per  hour.  These  facts  support  the 
belief  that  the  liver  plays  the  same  inipoi-tant  i>art  in  "assimilating" 
levulose  as  with  other  sugars. 

Spontaneons  Alimeniarj)  LcvuJosuria,  i.  c,  the  appearance  of  lev- 
ulose in  the  urine  from  such  small  quantities  of  levulose  as  occur 
naturally  in  the  food,  has  been  demonstrated  in  eight  cases.  In  five 
of  these  levulose  apjiears  to  have  been  the  only  sugar  present.  These 
persons  showed  a  decreased  tolerance  for  ingested  levulose  and  ceased 
passing  the  sugar  when   the  diet  was  cai-bohydrate-free.     The  tend- 

31  Bauer,  Dent.  nied.  Wocli..  IftdS  (.3.)),  150."):  Roiss  u.  .Tohn,  Dcut.  Arcli.  f. 
klin.  Med.,  1012  (108),  1S7:  Roiibitscheck,  Dcut.  Areli.  f.  klin.  Mod.,  1012  (108), 
225;  Naiuiyn,  "Bcitriifio  7,ur  Lchre  von  Ikterus,  etc.",  Reichcrt-Duboisschcs  Archiv 
fiir  Anatomie,  18G0,  p.  570;  Schupffer,  Arch.  f.  exp.  Path.  u.  Pharm,  1873  (1),  73. 


656  DIABETES 

ency  of  tli()U<ziit  would  ])o  to  look  i'of  ihc  cause  of  such  phenomena 
in  a  disturbed  hepatic  function. 

It  is  interesting  to  note  that  of  the  above-mentioned  five  cases  of 
pure  levulosuria,  two  showed  a  lessened  tolerance  for  glucose,  and  one 
symptoms  of  dis])itnitarism,  one  developed  during  the  puerperium, 
and  one  had  an  endocarditis ;  i.e.,  four  out  of  five  had  evidence  of 
derangements  of  the  endocrinous  glands.  The  literature  has  been 
reviewed  and  a  case  reported  by  Strouse  and  Friedman. ^- 

Mixed  Levulosuria,  or  the  occurrence  of  levulose  along  with  glucose 
in  severe  cases  of  diabetes,  is  said  by  some  to  be  a  common  event.  In 
view  of  the  great  frequency  of  combined  liver  and  pancreatic  changes 
found  at  autopsy  in  diabetic  cases,  and  in  view  also  of  the  freciuent 
occurrence  in  diabetes  of  signs  which  point  to  disturbances  of  other 
glands  with  internal  secretion  besides  the  pancreas,  this  would  har- 
monize well  with  the  view  just  given. 

Spontaneous  or  Idiopathic  Levulosuria,  having  a  character  similar 
to  chronic  pentosuria,  and  running  a  steady  course  uninfiueneed  by 
diet,  has  been  reported  in  one  case  by  Rosin.  In  this  instance  the 
tolerance  for  glucose  was  also  diminished. 

POLYSACCHARIDES 

Closely  related  to  these  meliturias  are  the  forms  in  which  the  poly- 
saccharides,-— lactose,  maltose  and  saccharose, — are  the  sugars  con- 
cerned. 

Lactosuria: — "When  2  to  3  grams  of  lactose  per  kilogram  of  body 
weight  are  given  in  pure  form  by  mouth  to  a  healthy  adult  dog  or 
man — alimentary  lactosuria  generally  occurs.  Another  form  of  lacto- 
suria is  that  seen  in  lactating  women.  In  these  cases  the  lactose  gains 
access  to  the  general  circulation  from  milk  stasis  in  the  breast.  Yet 
another  form,  the  lactosuria  in  chihlren,  having  gastro-intestinal  dis- 
eases, has  its  origin  in  the  lactose  of  the  milk  or  artificial  food.  In 
these  cases  lactosuria  may  develop  after  the  ingestion  of  lactose,  in 
quantity  and  form  ^^  incapable  of  causing  it  in  a  healthy  child.  The 
tolerance  for  lactose  is  most  strikingly  decreased  in  so-called  "intoxi- 
cation" (Finkelstein)  in  which  lactos\iria  may  follow  ingestion  of 
0.4-0.5  g.  per  kilo  of  body  weight.  (Grosz  places  the  assimilation  limit 
for  healthy  sucklings  at  8.6  g.  per  kilo.)  This  might  be  explained  in 
two  or  more  ways.  The  lactase  in  the  bowel  might  be  deficient  and 
permit  unhydrolyzed  sugar  of  milk  to  accunudate  in  abnormal  con- 
centration in  the  lower  bowel,  and  then  be  absorbed  unsplit ;  or,  as 
seems  more  probable,  the  bowel  wall — because  of  ulcers  or  simjile  in- 
flammatory changes — might  become  abnormally  permeable.  The  in- 
1ra\'euous  tolerance  limit  for  lactose  approaches  zero.     During  pro- 

32  Arcli.  Till.  :\rorl.,  1012    (0).  f)!). 

^^  Pun-  iHnicous  sohitions  (if  siitrio-  dilVcr  in   tlio  rato  of  absorption   from  those 
ill   wliicli  till-  siijrar  is  iiicor])(ira(('(l  in  hctcrojrcncous  mixtures. 


GLYCOSURIAS  657 

loiiji'ed  iiitravi'iioiis  injections  of  lactose  into  dogs  at  tlie  rate  of  2  fjin. 
])er  kilo  j)er  hour,  lactose  was  excreted  at  the  rate  of  injection  diii-iiif? 
the  fourth  hour  and  the  following  four  hours.'" 

Leopold  and  Reusse  reported  that  when  1  gram  of  la(;tose  was 
injected  subcutaneously  into  a  dog  or  infant,  exactly  1  gram  reap- 
peared in  the  urine;  but  that  if  the  injections  were  made  daily,  the 
([uantity  excreted  fell  little  by  little  and  finally  became  zero.  Ilelm- 
liolz  and  Woodyatt  have  repeated  this  experiment  in  dogs,  and  found 
that  at  first  the  gram  injected  might  reappear  in  the  urine  as  stated. 
Sometimes,  however,  the  occurrence  of  an  increase  in  the  reducing 
power  of  the  urine  above  the  figure  representing  1  gram  of  lactose 
was  noted.  This  suggested  a  splitting  of  the  lactose  into  glucose 
and  galactose.  Nor  could  they  obtain  more  than  a  temporary  disap- 
pearance of  the  sugar  following  subsequent  injections,  even  when 
carried  on  for  weeks — such  as  Leopold  and  Reusse  reported.  The 
point  of  chief  interest  in  these  experiments  is  that  the  apparently  in- 
creased hydrolysis  of  lactose  developing  with  successive  doses  resem- 
bles a  reaction  of  immunity,  with  a  substance  of  known  chemical 
composition  as  the  antigen.  But  it  is  possible  that  the  successive  in- 
jections simply  result  in  a  lessened  excretion  of  the  lactose  by  the  kid- 
ney's. Abderhalden  and  his  co-workers  reported  that  the  serum  of 
animals  similarly  treated  develops  an  increased  power  to  split  the  di- 
saccharide  employed,  as  determined  by  means  of  the  polariscope. 
These  experiments  were  paralleled  with  cane  sugar  (saccharose)  and 
with  di-,  tri-,  and  higher  peptids.  Other  observers  have  failed  to  cor- 
roborate these  findings. 

Saccharosuria  (cane  sugar  in  the  urine)  occurs  under  conditions 
quite  similar  to  those  mentioned  for  lactose,  except  that  there  is  no 
saccharosuria  corresponding  to  the  lactosuria  of  women. 

Maltosuria  has  often  been  reported,  but  the  chemical  detection  of 
this  sugar  is  uncertain. 

Other  polysaccharoses,  such  as  isomaltose,  glycogen,  etc.,  have  been 
thought  by  some  writers,  to  occur  in  the  urine. 

GLYCOSURIAS 

Glucose  is  the  sugar  which  enters  into  the  normal  glycogen  and 
forms  the  bulk  of  the  body  sugar.  Glycosurias  are  naturally  the 
most  important  of  the  meliturias. 

(1)  Alimentary  glycosuria,  e  saccharo.  Not  infrequently  it  is  im- 
possible to  make  a  healthy  man  eat  and  retain  sufficient  glucose  to 
cause  glycosuria,  and  it  would  be  hard  to  define  an  increased  glucose 
tolerance.  This  statement  is  corroborated  by  the  recent  studies  of 
Taylor  and  Hutton  ^■'  on  man.  Li  dogs  weighing  10  kilos  the  maxi- 
mum rate  of  glucose  absorption  is  apparently  reached  with  doses  of 

34  Unpublished  experiments  hv  W.  D.  Sansum. 

35  Jour.  Biol.  Cliem.,  lOlG   (25),  173. 

42 


658  DIABETES 

50  grams  and  ])('i-liaps  loss.  Larger  doses  do  not  further  increase  the 
rate  of  absorption.  This  rate  may  be  1.8  gram  per  kilo  per  hour.  If 
with  this  rate  of  absorption  the  rate  of  utilization  in  the  bowel  wall 
and  liver  is  0.9  gram  per  kilo  per  hour,  or  less,  glucose  will  enter  the 
systemic  blood  at  the  rate  of  0.9  gm.  per  kilo  per  hour,  or  more,  and 
this  will  normally  cause  glycosuria.  The  physiological  state  of  the 
liver  and  the  rate  of  sugar  absorption  are  factors  of  chief  importance 
in  determining  alimentary  glycosuria.  Glycosuria  following  the  in- 
gestion of  starch  alone — alimentary  gljjcosuria  ex  dmylo  is  said  not  to 
occur  in  healtliy  individuals. 

(2)  Glycosurias  which  depend  upon  the  discharge  of  sugar  from 
stored  glycogen.  These  may  be  due  to  the  action  of  (a)  nerves,  (b) 
drugs,  (c)  the  so-called  internal  secretions. 

(a)    Claude    Bernard's   piqure,    or   puncture    of    the    floor    of   the 
fourth  ventricle  between  the  points  of  origin  of  the  eighth  and  tenth 
pairs  of  nerves,  causes  a  glycosuria  which  ceases  when  the  glycogen 
of  the  liver  is  reduced  to  a  low  percentage.     Following  this  operation 
the  l)lood  is  found  to  contain  an  excess  of  sugar  (hyperglycemia")   to 
which  the  glycosuria  is  immediately  due.     If  the  vagus  nerve  is  cut 
stimidation  of  the  central  end  has  a  similar  effect,  so  that  the  vagus 
is  said  to  carry  the  afferent  impulse  to  the  center  in  the  calamus 
scriptorius.     By   severing   different   portions   of   the   nervous   system 
and  stimulating  the  cut  surfaces,  the  path  of  the  efferent  impulse  has 
been  traced  from  the  glj^cogenic  center  through  the  cord  to  the  upper 
thoracic  spinal  roots,  by  the  rami  commimicantes  to  the  inferior  cer- 
vical and  superior  thoracic  ganglion,  thence  via  the  splanchnic  nerves 
to  the   liver.     This   center   and   nervous  arc   form   probably   an   im- 
portant link  in  the  mechanism  for  regulating  the  quantity  of  sugar 
in  the  blood.     Nervous  glycosurias  having   the   same   mechanism   as 
"la  piqure"  occur  in  a  great  variety  of  conditions  associated  with 
insult  to  the  nervous  system,   e.  g.,  commotio  cerebri,  brain   tumor, 
tabes,  meningitis,  severe  mental  shock,  etc.     How  a  splanchnic  impulse 
operates  to  cause  increased  hydrolysis  of  glycogen  is  unsettled.     Gly- 
cogen hydrolyzes  outside  the  body  under  the  influence  of  acids,  /".  e., 
of  n  ions  or  plus  charges  of  electricity,  and  a  nerve  impulse  might 
tlieoretically  operate  directly,  or,  as  McLood  has  suggested,  through 
an  increase  of  glycogenase  in  the  liver;  or  as  held  by  the  von  Noorden 
school,  by  causing  an  increased  secretion  of  einnephrine — since  ]ii(|ure 
glycosuria  is  said  not  to  occnr  in  animals  deprived  of  the  adrenals 
'^  flayer,  Kahn,  Nishi)   or  after  section  of  tlie  left  splanchnic  nerve, 
which  supi)lies  both  adrenals,      (b)    Similar  phenomena  occur  in  as- 
])hy.\ia   (carbonic  and  lactic  acid  accumulation),  and  when  acids  are 
directly  administered;  also  alter  the  administration  of  certain  drugs 
whose  effects,  including  lactic  and  carbonic  acid  accumulation  in  the 
l)ody  fluids,  closely  ])aT-allel  those  of  a  deficient  oxygen  suj^jily   (phos- 
pliofiis,     carbon     monoxide,     chlorofoT-m,     iiNdi-azinc,     arsenic,     etc.). 


rillA)l{IIIZI\    DIMiETES  659 

Certain  (itlicr  (lrii<is,  siu-h  as  ciu-an',  strycliiiiu,  etc.,  may  iiitcrfei'c 
with  res{)irat()rv  moveiiieiits  and  so  cause  glycosuria  by  secondary 
asphyxia;  althougli  other  drugs,  of  which  tliere  are  many,  may  op- 
erate to  cause  glycosui-ia  in  any  of  the  waj's  by  which  glycosuria  can 
be  produced. ■''*' 

(c)  The  ductless  gland  extracts  which  produce  glycosuria  include 
those  of  the  adrenal,  thyroid  and  hypophysis.  Epinephrine  has  been 
discussed  in  another  place,  and  the  reasons  are  there  developed  for  the 
belief  that  the  glycosuria  it  causes  is  due  to  a  mobilization  of  sugar 
from  glycogen,  which  leads  to  hyperglycemia.  Kinger  ^'  showed  that 
when  an  animal  is  fully  ])hlorhizinized  the  svd)cutaneous  injection  of 
epinephrine  causes  no  additional  output  of  sugai'  nor  alteration  of  the 
G  :  N  ratio,  a  fact  which  has  been  contirmed  by  Sansum  and  AA^'oodyatt 
— thus  proving  that  epinephrine  has  no  power  to  intensify  a  diabetes 
which  is  already  at  the  point  which  is  called  complete.  Lusk  ^**  also 
showed  by  respiration  experiments  the  correctness  of  this  interpreta- 
tion. Eppinger,  Falta  and  Rudinger  ^^  stated  that  epinephrine  inten- 
sifies pancreas  diabetes,  and  used  this  observation  in  support  of  their 
idea  that  epinephrine,  like  thyroid  extract,  exerts  in  the  liver  a  sugar- 
mobilizing  and  sugar-building  etfect,  antagonistic  to  the  action  of  the 
pancreas,  which,  according  to  the  doctrine  of  the  von  Noorden  school, 
cheeks  the  formation  of  sugar  from  glycogen  and  also  from  protein 
and  fat.  But  in  their  work  there  has  been  no  adequate  proof  that  be- 
fore giving  the  epinephrine  the  pancreas  diabetes  was  as  complete  as  a 
pancreas  diabetes  can  be,  or  that  the  increased  intensity  of  the  dia- 
betes was  any  greater  than  could  have  been  explained  by  a  discharge 
of  sugar  from  glycogen.  The  power  of  pituitary  extracts  to  produce 
glycosuria  is  likewise  aseribable  to  their  effects  on  glycogen. 

PHLORHIZIN  DIABETES^" 

Phlorhizin  was  obtained  by  alcoholic  extraction  of  the  bark  and 
roots  of  apple,  pear,  plum  and  cherry  trees  by  L.  de  Koninck  in 
1835.  Its  glucosidic  character  was  established  by  Stas,  who  found 
that  it  could  be  split  into  glucose  ("phlorose")  and  a  substance 
(phloretin)  which  by  acid  hydrolysis  yielded  i)hloroglucin  and  an 
acid  (phloretinic  acid).  It  was  not  until  1886  that  von  Mering 
published  his  first  experiments  upon  its  physiologic  action. 

30  The  production  of  <jlycosiiria  by  a  piven  clni<r  should  not  be  confused  with 
an  excretion  of  yiaired  <rlycuronic  acid  compoiuids.  sucli  as  occurs  after  the 
administration  of  many  aldehydes,  ketones,  alcohols  and  phenols.  The  re- 
ducino-  power  in  these  cases  is  not  due  to  glucose  but  to  its  oxidation  product, 
COOH— ( CHOII ) ,— -COH. 

37  Jour.  Kxper.  INIed.,  1010  (12),  105. 

3s  Arch.  Int.  Med.,  1014  (13).  67.3. 

30  Zeit.  f.  klin.  Med.,  lOOS  (fifi).  1:  1900  (671,  3R0. 

4"  For  a  treatise  of  the  wliole  subject  of  phlorliizin  glycosuria,  with  bibliography, 
see  the  monograph  by  Lusk  (Phlorhizin  Glykosurio,  Ergeb.  der  Physiol.,  1012 
( 13 ) ,  315 ) ,  free  use  of  which  has  been  made  in  the  following. 


660  DIABETES 

While  phlorhizin  causes  glycosuria  when  taken  by  mouth,  its  great- 
est effect  is  obtained  by  subcutaneous  injection.  One  gram  of 
phlorhizin  triturated  in  5  to  15  c.c.  of  olive  oil,  or  in  20  per 
cent,  alcohol,  and  injected  subcutaneously  once  every  24  hours, 
will  maintain  the  maximum  glycosuria  which  can  be  produced 
in  a  dog  of  10  kilogrammes.  Phlorhizin  is  mostly  (80-90  per  cent.) 
excreted  in  the  urine.  It  is  soluble  in  ether,  optically  active,  and  gives 
a  garnet  coloration  with  ferric  chloride,  so  that  it  interferes  with  the 
polariscopic  tests  for  yS-hydroxybutj-ric  acid  in  the  urine,  and  masks 
the  (jcrhardt  reaction  for  aceto-acetic  acid. 

Phlorhizin  causes  glycosuria  in  frogs  and  other  cold-blooded  ani- 
mals, as  well  as  in  warm-blooded  forms  in  general,  including  birds. 
That  geese  show  glycosuria  with  phlorhizin  (von  Mering,  Thiel)  is 
important,  because  birds  do  not  pass  sugar  in  the  urine  when  op- 
erations are  performed  upon  them  which  do  cause  a  definite  excess 
of  blood  sugar  (pancreatectomy,  ^Minkowski).  Phlorhizin  causes 
glycosuria  in  birds — hyperglycemi-a  does  not.  Hence  phlorhizin  does 
not  cause  glycosuria  hy  producing  hyperglycemia.  In  harmony  with 
this  syllogism  are  the  data  obtained  by  Minkowski,  Levene,  von 
Czylharz  and  Schlesinger,  Lewandowsky,  Lepine,  Porcher,  Junkers- 
dorf,  Erlandsen,  Frank  and  Isaac — all  of  whom  have  found  the  blood 
sugar  concentration  in  phlorhizinized  animals  low  (0.065  per  cent. ; 
0.072  per  cent.;  0.012  per  cent.,  etc.).  Conflicting  results  have  also 
been  published,  but  the  methods  employed  in  these  instances  have  not 
usually  been  beyond  criticism  (Pavy,  Biedle  and  Kolisch).  Even 
after  ligation  of  the  renal  vessels  or  bilateral  nephrectomy,  no  hyper- 
glycemia has  been  demonstrated  in  phlorizinized  animals,  whereas  if 
phlorhizin  acted  by  liberating  sugar  from  glycogen  reserves  in  the 
liver  and  elsewhere,  or  from  any  source  distant  from  the  kidneys, 
hyperglycemia  might  be  expected.  In  view  of  these  facts  von  Mering 
liimself  interpreted  the  action  of  phlorhizin  as  a  kidney  diabetes. 

Zuntz  injected  phlorhizin  directly  into  one  renal  artery  and  col- 
lected the  urine  from  each  kidney  separately.  The  kidney  on  the 
injected  side  almost  at  once  secreted  saccharine  urine,  and  the  other 
kidney  secreted  sugar  only  after  the  lapse  of  minutes.  This  experi- 
ment has  been  successfully  repeated  by  others,  and  seems  to  prove 
that  phlorhizin  can  cause  glycosuria  by  acting  directly  on  the  kid- 
neys. The  many  experiments  which  have  been  made  to  determine 
the  relative  blood  sugar  content  of  the  renal  artery  and  vein  during 
phlorhizin  glycosuria,  add  little  to  this  subject. 

The  (luostions  arise:  Are  the  kidney  cells  the  only  structures 
ichich  are  directly  affected  hy  phlorhizin?  and,  What  is  the  exuct 
nature  of  the  phlorhizin  effect? 

Levene  collected  the  bile  of  phlorhizinized  dogs  and  found  that  it 
exhibited  reducing  power  after  the  phlorhizin  injection,  but  not 
before.     Ray,  McDermott  and  Lusk  failed  to  find  similar  properties 


PHLOIiniZIN  DIABETES  661 

in  vomited  bile  from  phlorhizinized  dogs.  Brauer  repeated  Levene's 
work — using  a  different  method  in  that  lie  cleared  the  bile  with  lead 
ficetate  prior  to  making  the  sugar  tests,  and  then  foimd  no  reducing 
substance.  AVoodyatt  obtained  results  like  Levene's  but  found  later 
no  reaction  for  sugar  when  the  bile  was  cleared  in  the  way  Brauer 
recommended.  Still,  in  the  native  state  it  yields  characteristic  crys- 
tals of  an  osazon,  and  ferments  with  yeast  after,  but  not  before 
phloi-hizinization.  Karl  rJrnbe  perfused  tortoise  livers  with  salt 
solution  containing  phlorhizin  and  was  able  to  cause  more  rapid 
deglycogenation  than  when  the  same  salt  solution  minus  phlorhizin 
was  used  in  control  experiments.  Now  Ray,  McDermott  and  Lusk, 
in  working  with  bile  which  had  been  in  the  alimentary  tract,  used 
material  that  had  had  time  to  lose  its  sugar  by  resorption.  Brauer 's 
clearing  method  may  take  out  a  trace  of  sugar  even  if  present  origi- 
nally, and  it  must  be  said  that  there  is  some  evidence  favoring  the 
idea  that  phlorhizin  acts  in  the  liver,  although  much  less  strongly 
than  in  the  kidneys.  Attempts  have  also  been  made  to  demonstrate 
a  direct  action  of  phlorhizin  on  the  mammary  (Cornevins)  and  sweat 
glands  (Delmare).  Cornevins'  positive  findings  were  not  confirmed 
by  Cremer  and  Porcher,  whereas  Delmare 's  work  has  not  been  re- 
peated. But  R.  Pearce,  working  with  a  blood-sugar  method,  found  an 
increase  of  sugar  in  the  pancreatic  juice,  and  Underbill  has  brought 
further  evidence  in  support  of  a  general  action,  il.  H.  Fischer  had 
some  nephrectomized  frogs,  which  are  able  to  live  indefinitely  in  water 
because  they  excrete  through  the  skin.  With  the  writer  some  of 
these  frogs  were  injected  with  phlorhizin  into  the  dorsal  lymph  sac, 
and  sugar  was  found  next  day  in  the  water  in  which  the  frogs  were, 
but  not  in  the  water  occupied  by  control  frogs.  The  possible  origin 
of  this  sugar  in  the  slime  makes  it  desirable  to  repeat  this  crucial  ex- 
periment. Although  the  view  most  commonly  held  is  that  phlorhizin 
acts  specifically  and  exclusively  on  the  kidney  cells  this  has  never 
been  proved,  and  there  is  much  to  suggest  a  general  cell  effect  ex- 
hibited most  strikingly  in  the  kidney. 

Regarding  the  fundamental  nature  of  the  action  of  phlorhizin, 
nothing  satisfactory  has  been  evolved.  Minkowski  suggested  that 
phloretin  and  sugar  are  split  apart  in  the  kidney  epithelium,  and 
that  the  sugar  is  then  excreted  while  the  phloretin  is  retained  in 
the  body.  The  retained  phloretin  then  takes  up  a  new  molecule  of 
glucose  from  the  blood  to  reform  phlorhizin, — which  in  the  kidney 
is  again  split,  etc.  (vehicle  theory).  Zuntz  has  determined  with 
a  given  minute  dose  of  phlorhizin  how  much  sugar  can  be  elimi- 
nated in  a  given  time  by  one  kidney ;  then,  figuring  what  weight  of 
phlorhizin  is  in  the  kidney,  and  how  much  sugar  comes  out  of  the 
kidney,  he  reckons  how  frequently  the  synthesis  and  hydrolysis  of 
phlorhizin  would  have  to  occur.  He  makes  it  26  times  per  minute, 
which  he  deems  too  fast  to  be  probable,  but  in  view  of  the  work 


662  DIA  BETES 

wliieh  can  be  accomplished  by  traces  of  organic  and  inorganic  car- 
riers (catalyzers,  enzymes),  this  criticism  is  not  convincing. 

Whatever  the  action  of  phlorhizin  niaij  prove  ultimately  to  he, 
this  action,  finds  its  chief  or  final  expression  in  the  cells  of  the  kid- 
ney, and  there  leads  to  a.  disturbance  of  equilibrium,  whereby  the 
relative  blood  sugar  and  urinarij  sugar  concentrations  are  altered 
in  favor  of  the  urine.  The  blood  sugar  nnist  be  in  equilibrium 
with  the  sugar  content  of  the  various  cells,  and  this  with  the  sources 
(glycogen  and  protein)  from  which  the  sugar  comes.  The  sugar 
of  the  entire  body  may  be  conceived  of  as  a  gas  exerting  its  partial 
pressure  in  every  cell  and  body  fluid, — here  more  dense,  there  less  so, 
depending  upoii  local  physico-chemical  conditions,  but  nevertheless 
everywhere  in  communication.  Phlorhizin  acting  in  the  kidneys,  and 
regardless  of  a  possible  action  elsewhere,  creates  a  void  into  which  the 
blood  sugar  flows,  and  into  which  secondarily,  as  into  a  vortex,  sugar 
flows  from  all  the  sources  of  the  body. 

Metabolic  Phenomena. — When  a  fasting  dog  is  kei)t  continuously 
under  the  maximum  effects  of  phlorhizin,  there  is  at  first  a  verj-  great 
glycosuria  while  the  urinary  nitrogen  remains  low.  The  ratio  of  the 
urinary  glucose  to  the  urinary  nitrogen  (G  :  N  ratio)  may  be  as  high 
as  10  or  15  to  1,  or  higher.  If  such  a  dog  is  killed  the  liver  is  found 
to  have  a  normal  appearance  and  to  contain  glycogen.  As  time  goes 
on  the  rate  of  glucose  excretion  falls  and  the  nitrogen  tends  to  in- 
crease, until  after  two  or  three  days  the  G  :  N  ratio  is  about  3.65  to  1, 
as  shown  by  Lusk.  Then  for  12  to  24  hours  it  may  remain  constant. 
It  sometimes  happens  that  the  ratio  falls  to  2.8  or  some  point  between 
3.65  and  2.8  before  constancy  is  established.  It  then  proceeds  at  this 
lower  level  instead  of  3.65.  If  a  dog  is  killed  at  about  the  time  con- 
stancy is  attained,  or  somewhat  sooner,  the  liver  may  be  found  in  a 
state  of  fatty  infiltration  with  the  glycogen  low  but  not  absent.  In 
later  stages  the  excessive  fat  in  the  liver  again  disappears.  There  is 
then  first  a  rapid  loss  of  glucose  and  a  consequent  melting  away  of 
glycogen.  To  compensate  for  the  falling  out  of  the  carbohydrate 
from  the  nieta])olism  there  is  an  increased  breakdown  of  i)rotein  and 
a  rapid  mo])i]i/ation  of  fat,  finding  tempoi-ary  expression  in  a  fatty 
infiltration  of  the  liver.  But  as  the  fat  resei-vcs  rnn  low  tlio  fat  de- 
posited in  the  liver  is  utilized.  Coincident  with  the  ])artial  exhaustion 
of  the  carbohydrate  reserves  of  the  body  and  the  increased  fat  and 
protein  metabolism,  acetoacetic  and  ^-hydroxy  butyi-ic  acids  begin  to 
appear  in  the  urine,  and  since  they  are  excreted  jiartly  in  the  form  of 
the  ammonium  sails  flic  urinary  ammonia  is  also  increased.  These 
acids  arise  from  lower  fatty  acids  having  an  even  inimber  of  carbon 
atoms  in  the  chain,' and  from  certain  amino-acids.  whenever  the  mix- 
ture of  fatty  acids  and  L'lncosc  actually  nictabolizini;-  is  loo  I'icli  in  1lie 
former  in  comparison  willi  llic  lalio-. 

lTovv('\('r,  such  animals  ai'c  not   U-i.'^  of  <ih-co<i'en.     If  thev  are  sub- 


l'lll.nh'l////\   DIAHKTI.s  663 

jc'cted  to  some  treatnuMit  wliii-h  has  a  strong-  j^lycogen  niohili/iiij,'  ef- 
fect the  glycosuria  may  be  made  to  rise  temporarily,  just  as  though 
the  dog  had  been  given  a  dose  of  sugar.  Thus,  exposure  to  cold  suf- 
ficient to  cause  shiverinji:,  the  administration  of  ('i)iiu'phrinc.  or  an 
ether  or  nitrous  oxide  narcosis,  injection  of  acid  (and  various  other 
toxic  substances  capable  of  producing  tissue  asphyxia  and  acidosis),  all 
may  increase  the  urinary  glucose  without  increasing  the  nitrogen,  and 
thus  cause  an  increased  G  :N  ratio.  P>ut  if  the  exposure  to  cold  is 
long  and  intense  enough  a  time  conies  when  it  ceases  to  have  this  ett'ect, 
and  if  epinephrine  is  given  subcutaneously  in  the  dosage  of  about  0.4 
mg.  per  kg.  of  body  weight  once  ever}-  three  hours  there  is  for  a  tinie 
a  heavy  increase  of  the  glucose  output,  but  this  becomes  less  and  less 
until  after  6  or  8  doses  the  ratio  becomes  constant  again,  regardless  of 
whether  epincj)hrine  is  given  or  not.  In  such  dogs  neither  cold  nor 
narcosis  nor  other  toxic  effects  will  increase  the  output  of  glucose, 
and  analyses  of  the  liver  and  muscles  reveal  no  glycogen.  In  a  long 
series  of  dogs  so  treated  Sansum  and  the  writer  have  not  encountered 
ratios  above  3.2  to  1,  and  the  2.8  ratio  recurs  frequently. 

Since  the  glycogen  is  gone  and  the  dog  is  fasting,  the  sugar  which 
continues  to  appear  in  the  urine  must  have  its  origin  in  body  fat  or 
protein,  or  both. 

Sugar  from  Fat. — If  such  a  dog  be  given  large  cpiantities  of  fat 
in  the  diet  no  change  occurs  in  the  G  :  N  ratio,  nor  any  increase  in 
the  glycosuria,  except  such  as  may  be  ascribed  to  the  glycerol  of  the 
fat  (Lusk).  On  the  other  hand,  propionic  acid,  according  to  Ringer, 
may  cause  a  rise  in  the  sugar  excretion  and  a  corresponding  rise  in 
the  G  :  N  ratio.*^  From  this  it  is  concluded  that  the  fats  of  the  food 
do  not  as  a  rule  form  sugar  in  the  body,  although  sugar  formation 
from  at  least  one  lower  fatty  acid  is  possible  in  view  of  Ringer's 
experiment. 

Von  Noorden  and  Falta  and  their  associates  have  regarded  sugar 
formation  from  fat  as  a  regular  normal  ]>henomenon,  because  in  dia- 
betes melitus  they  believe  that  high  ratios  occur  wliich  make  this 
view  necessary. 

Sugar  from  Protein. — If  instead  of  fat,  ]irotein  be  given  to  the 
dog  above  mentioned,  there  occurs  an  absolute  rise  in  the  sugar  of 
the  uriiu^  and  a  corres])onding  rise  in  the  nitrogen,  hut  the  G:X 
ratio  remains  constant.  Following  a  meat  feeding  there  may  be  fluc- 
tuations of  the  ratio  during  short  periods,  but  this  statement  generally 
holds  if  the  time  of  observation  is  12  to  24  hours.  These  facts  have 
led  to  the  conclusion  that  when  in  a  fasting,  fully  phlorhizinized  ani- 
mal, or  one  fed  on  meat  and  fat  alone,  a  constant  (r  :  X  ratio  of  3.65  :  1 
is  seen ;  this  means  that  the  glucose  and  the  nitrogen  are  coming  from 
one  and  the  same  source,  viz.,  protein.     A  cram  of  uitrotren  corresponds 

*^  The  do<rs  used  l)y  Uiiisrer  wore  not  free  of  glycogen  and  possibly  the  extra  sugar 
did  not  arise  from  the  acid  given. 


664  DIABETES 

to  6.25  grams  protein,  and  if  for  each  6.25  grams  pi*otein  metabolized  as 
indicated  by  the  N  in  the  urine,  3.65  grams  glucose  are  excreted,  then 
58  per  cent,  of  the  protein  metabofi/.ed  is  converted  into  glucose  and 
so  excreted.  In  like  manner  the  2.8  : 1  ratio  would  indicate  a  45  per 
cent,  conversion.  A  percentage  above  58  has  not  been  satisfactorily 
proven  to  occur.  If  to  the  fully  phlorhizinized  dog  a  definite  quan- 
tity of  glucose,  galactose,  starch  or  other  assimilable  form  of  carbo- 
hj'drate  is  given,  this  may  under  favorable  circumstances  be  excreted 
quantitatively  in  the  urine  as  glucose,  and  the  ratio  of  G  :  N  will  rise. 
The  sugar  which  appears  in  the  urine  under  such  circumstances  over 
and  above  that  represented  by  N  X  Gr  :  N  has  been  called  "extra 
sugar"  by  Lusk. 

If  all  the  carbon  contained  in  protein  were  converted  into  glu- 
cose, and  all  this  excreted  together  with  the  nitrogen,  the  G  :  N  ratio 
would  be  8.25  : 1.  A  higher  ratio  than  this  would  necessarily  mean 
that  sugar  was  coming  from  some  source  other  than  protein,  or  that  all 
of  the  N  was  not  appearing  in  the  urine,  some  being  retained  in  the 
bod}'.  If  the  liver  were  free  from  glycogen  and  no  carbohydrate 
were  eaten,  such  a  high  ratio  would  speak  in  favor  of  sugar  forma- 
tion from  fat.  Falta  reports  having  seen  cases  of  diabetes  in  which 
this  occurred,  but  in  human  cases  it  is  difficult  to  be  sure  of  the 
absence  of  glycogen  and  food  carbohydrate  ;  moreover,  such  high  ratios, 
unless  too  long  continued,  might  imply  retention  of  nitrogen.  Thus, 
in  thyreopriva,  a  nitrogenous  substance  is  retained  in  the  body 
^myxoedema  fluid)  ;  if  this  substance  contains  more  N  than  the  total 
weight  divided  by  6.25,  a  portion  of  protein  must  be  metabolized, 
having  a  lower  percentage  of  N.  This  latter  portion  could  give  a 
higher  G  :  N  ratio  than  normal  protein. 

Sugar  from  Other  Substances. — A  large  number  of  other  substances 
when  administered  to  phlorhizinized  dogs  are  capable  of  increasing 
the  output  of  sugar.  Of  importance  in  tliis  connection  are  certain  of 
the  amino  acids,  viz. :  glycine,  alanine,  aspartic  and  glutamic  acids, 
and  arginine.  Others,  such  as  leucine,  tyrosine  and  phenyl  alanine 
do  not  form  sugar  in  the  body  but  increase  the  output  of  the  acetone 
substances.  Tlie  sugar-forming  power  of  protein  is  doubtless  due  to 
its  content  of  the  former  group  of  amino  acids.^-  Lactic  acid  and 
glycerol  are  also  among  the  sugar  formers. 

The  chief  interest  in  phlorlii/in  diabetes  lies  in  tlie  opportunities 
it  offers  of  studying  the  charac'ter  of  the  intermediate  metal)olism 
minus  that  of  sugar,  and  so  of  studying  sugar  metabolism.  Another 
interest  might  be  found  were  the  physiologic  effects  of  this  glucoside 
in  animals  interpreted  with  relationsliip  to  its  normal  role  in  plant 
physiology. 

42  See  Dakin,  Jour.  Riol.  Clicin.,  1913  (14),  155. 


PANCREAS  DIABETES  AND  DIABETES  MELITUS  665 

PANCREAS  DIABETES  AND  DIABETES  MELITUS 

Historical. — In  1788  Cowley  reported  atrophy  and  stone  of  the 
pancreas  in  a  case  of  diabetes.  The  coincidence  of  diabetic  symp- 
toms and  lesions  of  tlie  pancreas  was  furtlier  studied  by  Bright, 
Lloyd  and  Elliotson  (1833).  It  was  Bouchardat "  who  first  defi- 
nitely formulated  the  belief  that  pancreatic  disease  was  the  cause  of 
diabetes  melitus,  but  his  views  were  uncongenial  to  the  clinicians  of 
his  time  and  it  remained  for  von  Mering  and  ^Minkowski  ^^  (1889)  to 
prove  that  complete  pancreatectomy  leads  invariably  to  the  devel- 
opment of  a  severe  diabetes.  This  applies  not  only  to  dogs  but  to 
cats,  rabbits,  pigs  (Minkowski),  tortoises,*'^  frogs,*'^  eels,**'  and  other 
animals. 

Effects  of  Pancreas  Extirpation. — The  gh'cosnria  begins  soon  after 
the  operation  and  increases  in  intensity.  It  persists  in  spite  of  a 
non-carbohydrate  diet  long  after  the  glycogen  reservoirs  in  the  liver 
and  muscles  have  become  greatly  impoverished  (to  0.1-0.2  per  cent, 
in  the  liver),  but,  like  the  human  disease,  it  usually  ceases  during 
a  fast  or  may  disappear  just  before  death.*"  The  glycosuria  may  be 
accompanied  by  an  excretion  of  the  acetone  bodies, — acetone,  aceto- 
acetic  and  )8-hydroxybutyric  acids.  In  fact,  the  metabolic  changes 
secondary  to  this  operation  closely  parallel  those  found  in  the  humai? 
disease,  with  certain  difit'erences  which  perhaps  are  ascribable  to  species 
or  to  the  fact  that  in  the  experimental  diabetes  digestion  is  altered  by 
absence  of  the  pancreatic  juice,  etc.  Although  ^Minkowski 's  work  was 
assailed  from  many  quarters,  the  following  points  have  become  firmly 
established  by  frequent  repetition.  (1)  Complete  removal  of  the 
pancreas  causes  a  true  diabetes  (as  above)  ;  (2)  Ligation  or  oblitera- 
tion of  the  duct  (or  ducts)  of  Wirsung,  no  matter  how  scrupulously 
carried  out,  has  no  such  effect;  (3)  If  about  one-fifth  of  the  pancreas 
with  its  arterial  supply  be  separated  from  the  rest  of  the  gland,  this 
fifth  may  be  implanted  extraperitoneally  at  a  distance  from  the  origi- 
nal site.  No  diabetes  results  from  this  operation,  or  at  most  only  a 
transient  glycosuria.  Now  if  the  main  body  of  the  pancreas  be  fully 
extirpated  with  ducts,  nerves  and  bloodvessels,  still  only  a  transient 
glycosuria  or  none  at  all  develops.  At  this  stage  all  possible  damage 
to  nerves  and  external  secretion  has  been  inflicted  and  proven  in- 
capable of  causing  diabetes.  (4)  In  the  course  of  weeks  the  graft 
atrophies  (Sandmeyer's  experiment),  and  then  a  persistent  glycosuria 
supervenes;  or  the  encapsulated  fragment  which  has  been  placed  in 
an  accessible  place  under  the  skin  may  be  extirpated,  in  which  case 

43  "De  la  Glvcosurie,  etc.,"  II  edit.,  Paris,  1883.     Cited  from  Nauiivn. 

44  Arch,  fiir  exp.  Path.  u.  Pharm.,  1889  (26),  371;  185)3  (31).  85. 
45Aldehoff,  G.   Zcit.  f.   Biol.,   1891-2    (28),  293:   Velich,  Wien.  Med.  Zeitung., 

1895    (40),  502:  :^rarcuse  W..  Zeit.  f.  klin.  Med.,  1894    (26),  22.5. 

46  Capparelli,  Biol.  Zentralbl.,  1893    (13),  495. 

47  This  statement,  based  on  experimental  work,  appears  in  the  2d  (1914)  edition 
of  this  book. 


666  DIABETES 

within  a  few  hours  a  severe  diabetes  ensues.  (5)  There  is  no  other 
organ  in  the  body  extirpation  of  which  has  any  similar  effect,  nor 
(except  for  pldorhizinization),  is  tliere  any  known  means  of  experi- 
mentally producing  a  true  diabetes  without  injury  to  the  pancreas. 
(6)  No  toxic  substance  derived  from  the  bod}^  of  diabetic  individuals, 
man  or  animal,  lias  been  found  which  is  capable  of  causing  diabetes  in 
a  second  animal.  These  facts  lead  to  the  conclusion  {reached  hy  Min- 
Jcowski)  that  pancreatic  tissue  provides  "a  something,"  separate  from 
the  pancreatic  juice,  {internal  secretion  of  the  pancreas),  the  lack  of 
ichich  is  responsible  for  the  si/mptoms  of  diabetes. 

Islet  Theory:  IMorpliologically  the  pancreas  may  be  regarded  as  stroma, 
ducts,  acini  and  islands  of  Langerhans.  It  has  been  proposed,  notably  by  Opie  *^ 
in  this  country,  that  the  antidiabetic  internal  secretion  of  the  pancreas  is  elabo- 
rated by  islet  cells.  This  view  finds  support  in  the  followinof  facts:  (1)  In 
diabetes  melitus  the  islets  are  frequently  foimd  in  a  state  of  hydropic  or  hyaline 
degeneration,  while  tlie  remaining  organ  may  appear  normal. *o  (2)  Cancer,  pan- 
creatitis and  tiie  experimental  injection  of  caustics  into  the  ducts  very  frequently 
spare  tlie  islets  and  fail  to  cause  diabetes.  (3)  It  is  claimed  that  in  ])ancreatic 
grafts,  such  as  described  above,  islet  cells  predominate,  while  acinus  cells  and 
ducts  disappear. 

(irafts  of  this  kind  consist  of  much  connective  tissue,  generally  more  or  less 
infiltrated  with  round  cells,  and  collections  of  epithelium.  Concerning  the  latter, 
remains  of  ducts  and  acini  are  usually  present  in  some  proportion,  and  there  axe 
also  epithelial  cell  masses  regarded  as  islets  on  morphological  groruids.  Differ- 
ences of  opinion  still  exist  as  to  the  relative  proportion  of  the  diiTerent  epitlielial 
elements.  Lombroso,'>o  whose  exhaustive  monograph  reviews  the  literature  to  1910, 
concludes  tliat  the  internal  function  of  the  pancreas  is  not  monopolized  by  islet 
cells.  Bensley  si  developed  intra-vital  staining  methods  which,  for  tlie  first 
time,  made  possible  the  sure  differentiation  of  islet  cells  from  duct  or  acinus 
epithelium  witliout  reference  to  form  or  arrangement,  and  appears  to  have  proved 
tliat  these  cells  are  regenerated  from  duct  epithelivun.  lie  also  showed  the  great 
normal  variations  in  size  and  number  of  islets  in  different  individuals  (guinea 
pigs).  His  study  explains  certain  of  the  discrepancies  which  occur  in  tlie  litera- 
ture, es])ecially  in  tlie  estimation  of  the  quantity  of  islet  tissue  in  pancreatic  rests, 
grafts,  etc.  Recently  Allen  52  has  reported  that  when  proper  sized  fragments  of 
pancreas,  in  connecticm  with  the  ducts,  are  left  in  situ,  and  the  remainder  of  the 
gland  is  removed,  the  subsequent  development  of  severe  diabetes  may  be  coincident 
with  disappearance  of  islet  tissue  while  acinus  cells  and  ducts  are  unaffected. 
This  operation,  according  to  Allen,  is  eminently  satisfactory  for  producing  ex- 
perimental diabetes  without  infection  and  without  loss  of  the  external  secretions. 

The  Nature  of  the  Internal  Secretion  of  the  Pancreas. — Direct  evi- 
dence on  this  subject  is  lacking.  Such  a  secretion  has  never  been 
isolated.  Even  the  experiments  made  with  tlie  feeding  of  fresh  pan- 
creas and  with  extracts  of  the  gland  have  led  to  no  definite  advance. 
R<'ports  of  iniiirovement  following  the  administration  of  any  sub- 
stance in  dialx'tes  are  worthless  unless  accompanied  by  proof  of  the 
constancy  of  the  diet,  of  th(>  amount  of  work  performed,  and  of  other 
factors  wliich  are  known   to  influence  the  course  of  diabetes.     Some 

48  "Diseases  of  the  Pancreas,"'  LipjiincoH  >!c  Co.,  1910. 
40  See  llomans,  .Tour.  MvA.  Kes..  1014    (.30),  49. 
noErgeb.  der  Plivsiol.,  1910   (10),  1. 
•'.lAm.  Jour,  of  Anat.,  1911    (12),  297. 
52  Gl.vcosuria  and  Diabetes,  Bostojj,  1913. 


/'.I  \r/,'/;.iN  hiMii:Ti:s  .i\/>  m  \iu:ti:s  \ii:i.iTUi^  667 

jrlinimcr  of  success  ajjpcarcd  to  liavc  altcndcd  the  iiilravcuous  use  of 
an  extract  made  1»\  Zurl/.ci-.  ■  ■  although  delclci'ioiis  hy-cffects  occurred, 
and  the  apparent  ini])rovenient  could  have  been  due  wliolly  to  reten- 
tion. According  to  lledon  and  Drennan,  amelioration  of  tiio  severity 
of  pancreas  diabetes  as  evidenced  by  a  dimimition  of  glycosuria  has 
followed  the  transfusion  of  blood  from  a  healthy  animal  or  the  injec- 
tion of  fresh  detibrinated  blood,  and  Forschbach.  working  with  a 
])arabiosis  (or  two  animals  so  joined  by  oi)erative  means  that  pei-ma- 
nent  intermingling  of  their  blood  occurs)  performed  pancreatectomy 
ill  one  of  the  animals  without  producing  diabetes  in  either;  from  which 
it  might  seem  that  the  internal  secretion  was  carried  by  the  blood. 
Ill  harmony  with  these  results  were  the  investigations  of  Knowlton 
and  Starling,''*  who  found  that  an  isolated  beating  heart  taken  from 
a  depancreatized  animal  (cat)  was  capable  of  removing  less  sugar 
from  the  blood  used  as  a  perfusion  medium  than  are  hearts  of  normal 
animals,  but  these  latter  experiments  have  not  been  confirmed  and  are 
subject  to  criticism.  Tn  most  of  the  transfusion  experiments  re- 
ported the  standardization  of  the  metabolism  prior  to  giving  the  fresh 
blood  has  not  been  such  as  to  make  the  results  certain.  Carlson  and 
Drennan  found  that  pancreatectomy  in  a  pregnant  animal  near  term 
might  fail  to  cause  diabetes,  but  that  diabetes  developed  at  once  fol- 
lowing delivery.  This  could  be  explained  on  the  basis  that  an  in- 
ternal secretion  passed  from  fetus  to  mother,  or  that  sugar  failing  of 
utilization  in  the  mother  was  utilized  by  the  fetuses.  Kramer  and 
JNTurlin  failed  to  note  any  increase  of  the  respiratory  quotient  in  de- 
pancreatized  dogs  following  blood  transfusion,  and  Sansum  and 
Woodyatt  saw  no  improvement  following  transfusion  in  a  human 
case."" 

Symptoms. — Tn  the  absence  of  extracts  which  contain  the  active 
]iriiici]ile  in  measurable  fpiantity,  the  attention  must  be  turned  to  a 
more  detailed  study  of  the  effects  which  follow  its  lack.  Now  it  is 
well  known  that  in  diabetes  melitus  there  are  all  grades  of  severity. 
WTiai  follows  has  reference  onily  to  the  severest  cases — those  ivhich 
maif  hr  called  "complete  diahetes."  In  the  severest  cases  of  dia- 
betes, glycosuria  ])ersists  even  when  the  individual  subsists  on  a  fat- 
protein  diet,  and  after  the  glycogen  in  the  body  has  been  reduced  to 
a  mere  trace.  When  this  stage  has  been  reached,  and  provided  no 
carbohydrate  food  is  eaten,  it  is  found  that  tlie  total  glucose  in  the 
urine  bears  from  day  to  day  a  constant  ratio  to  the  total  nitrogen  in 
ihe  urine  as.already  described  for  })lilorhizin  dialietes.  This  "G  :N" 
ratio"  is  not  always  the  same.  Tn  depancreatized  dogs  nourished 
solely  on  fat  and  protein,  it  is  often  found,  as  Minkowski  first  recog- 
nized, at  2.8  :1,  and  in  human  diabetes  the  same  value  for  G  :  N  is 

•-••iZoit.  f.  oxp.  Pali)..  innS-n   (5),  .107. 

54  .Tom-,  of  Physiol.,  1913  (45),  140. 

55  Jour.  Amer.  Med.  Assoc,  1914   (62),  006  for  lit.  references. 


668  DIABETES 

sometimes  seen.  But,  as  in  phlorhizinized  dogs,  higher  ratios  may 
occur  in  the  human  disease. 

If  to  such  a  case  of  diabetes  as  this  we  give  by  mouth  40  grams  of 
glucose  there  may  appear  in  the  urine  close  to  40  grams  of  extra 
sugar.  Plainly  such  extra  sugar  has  escaped  utilization  of  any  kind. 
It  cannot  have  been  oxidized  or  converted  into  fat,  since  these  proc- 
esses are  irreversible,  although  it  might  have  existed  momentarily  in 
the  body  as  glycogen  or  other  isomer  of  glucose.  What  phase  in  the 
utilization  of  this  glucose  is  primaril}'  disturbed  is  another  question. 
To  say  that  40  grams  of  ingested  glucose  causes  the  appearance  of  40 
grams  of  extra  sugar  in  the  urine  does  not  prove  that  the  diabetic 
body  is  inherently  incapable  of  using  any  sugar  or  every  carbohydrate. 
It  might  still  be  capable  of  using  a  two,  three,  or  four  carbon  atom 
sugar,  some  other  member  of  the  group  of  32  hexoses,  or,  as  some  have 
it  (von  Noorden),  sugar  which  has  first  been  built  up  into  glycogen, 
etc.,  provided  these  substances  could  be  kept  from  undergoing  trans- 
formations into  the  non-utilizable  glucose.  As  a  rule,  however,  when 
other  sugars  are  fed  to  complete  diabetics,  they  are  transformed  into 
glucose  and  appear  as  such  in  the  urine.  This  phenomenon  has  much 
of  significance  for  the  general  theory  of  sugar  metabolism  and  is  an 
indication  of  the  nature  of  the  primary  disturbance  in  diabetes,  as  will 
now  be  shown. 

Theory  of  Diabetes. — What  sort  of  a  chemical  process  is  involved 
when  levulose,  for  example,  is  converted  in  the  body  into  glucose? 
As  already  stated  in  the  chemical  introduction,  the  reciprocal  trans- 
formations of  hexoses  one  into  another  in  the  alkaline  solution  in 
liiro  depend  upon  a  preliminary  ionization  of  the  sugars  followed  by 
salt  formation,  the  salts  then  undergoing  dissociation  which,  according 
to  ^lathews  and  IMichaelis,  is  still  purely  electrolytic  with  rearrange- 
ments of  the  anion ;  but  which,  according  to  Nef,  is  a  non-electrolytic 
dissociation  of  the  type  which  he  calls  methylene  dissociation.  Some 
form  of  dissociation  must  he  a  prelude  also  to  these  transformations  in 
the  hody.  This  view  is  logically  just  as  necessary  as  it  has  been  found 
to  be  for  the  organic  chemist,  and,  it  may  be  added,  that  for  the  oxida- 
tion of  sugars  as  well  as  for  their  polymerization  a  preliminary  dissoci- 
ation is  essential.  Now  since  the  diabetic  body  can  transpose  other 
sugars  into  glucose,  it  must  be  able  at  least  to  dissociate  the  former 
sugars  deeply  enough  for  this  process.  These  trans])ositions  are  ac- 
complished chiefly  in  the  portal  system  and  perhaj^s  in  other  places 
too,  but  certainly-  levulose  and  man}'  other  substances  can  be  made  in 
the  liver  into  glycogen,  whose  hj'drolysis  then  yields  glucose. 

The  degree  or  character  of  the  dissociation  necessarj''  for  reciprocal 
transfonnations  differs  from  that  which  is  a  necessary  prelude  to 
destructive  reactions  such  as  oxidation.  A  very  weak  alkali  suffices 
in  vitro  for  the  former,  while  for  the  latter  it  is  necessary  to  use  a 


THEORY  OF  DIABETES  669 

soinewliat  stronger  alkali  t'oiiceiitratioii. '"■  The  diabetit'  body  tlK-re- 
fore  behaves  as  tliouj^h  it  were  weakened  witli  respect  to  the  alkali  eou- 
ceiitratioiis  which  it  cau  bring  to  bear  on  sugars. 

As  far  back  as  1871,  Sehultzen  suggested  that  the  error  in  diabetes 
might  be  found  in  the  disability  of  the  body  to  dissociate  the  glucose 
juolecule  into  two  3-carbon  substances.-"  i^aunigarten  "'^  also  supported 
the  idea  of  a  "fermentative  splitting"  which  precedes  oxidation,  be- 
cause he  found  a  greater  percentage  utilization  of  certain  substances 
closely  allied  to  glucose  (such  as  gluconic  acid,  saccharic  acid,  mucic 
acid,  etc.),  than  of  glucose  itself;  whereas  gluconic  acid  and  glucose, 
for  example,  differ  only  in  that  the  sugar  has  an  aldehyde  group  where 
the  acid  has  carboxyl.  Similar  general  ideas  have  been  expressed 
from  time  to  time  by  others.  The  present  writer  has  urged  in  place 
of  the  vaguer  terms,  the  adoption  of  chemical  "dissociation"  in  the 
sense  which  is  rapidh'  finding  favor  in  the  field  of  pure  organic  chem- 
istry, notably  for  the  explanation  of  the  behavior  of  aldehydes,  ketones 
and  alcohols.'^'-'  There  can  be  no  doubt  that  the  dissociation  of  glucose 
in  the  body  is  a  normal  occurrence.  This  is  directly  and  conclusively 
shown  whenever  muscles  make  lactic  acid  (C.jHqO.J  out  of  glucose 
(CgHijOo),  since  in  this  process  no  chemical  phenomenon  is  involved 
save  cleavage  of  the  hexose  and  intramolecular  rearrangement.  The 
polymerization  of  sugar  into  glycogen  might  be  similarly  interpreted. 
Direct  proof  of  a  failure  of  glucose  dissociation  in  diabetes  has  not  yet 
been  brought,  although  its  absence  would  explain  all  the  metabolic 
phenomena  more  directly  and  simply  than  any  other  single  physi- 
ologic error  which  has  been  hypothecated.  It  is,  moreover,  a  tangible 
chemical  conception,  whereas  to  say  that  the  body  loses  its  power  to 
oxidize  sugar  or  to  "fix"  it  as  glycogen  is  merely  to  name  effects  in 
phj^siologic  terms.      (Cf.  Naunyn's  diszoamylie). 

It  might  be  assumed  that  all  sugars  upon  entering  certain  phases  of 
the  cells  (phases  especially  well  represented  in  liver  cells),  meet  con- 
ditions which  are  equivalent  to  those  met  in  a  weakly  alkaline  solution, 
favoring  reciprocal  transformations,  and,  as  A.  P.  Mathews  points 
out,  polymerization;  but  not  conditions  conducive  to  the  deeper  de- 
structive reactions.  That  is,  especially  in  the  liver,  there  may  be  the 
equivalent  of  dilute  alkali  for  all  sugars.  Glucose,  being  the  least 
dissociable,  represents  the  form  into  which  all  other  sugars  tend  to 
accumulate.  But  in  the  normal  body  a  special  glucolytic  enzyme 
Calkali  carrier  or  intensifier?)  destroys  glucose  selectively.  All  other 
sugars  must  become  glucose  before  destruction.  In  diabetes  the 
enzyme  necessary  for  the  deep  dissociation  of  glucose  is  lacking  or  in- 

50  See  Woodyatt,  Jour.  Biol.  Chom.,  1015  (20),  129. 

57  Zeit.  f.  oxp.  Path.  u.  Pliarm..  1005  (2) .  53. 

58  Glyceric  aldehyde  and  glycerol,  according  to  Sehultzen. 

59  Cf.  Xef,  loc.  cit.,  and  Stieglitz,  Qualitative  Chemical  Analysis.  New  York,  1912, 
I,  pp.  289-292. 


670  DfAnKTES 

active.  Tlie  recent  studies  of  Murliii,  Kramer,'  Sweet  ami  Karver, 
show  that  alkali  administration  (NaoCO;,)  may  increase  glucose  utili- 
zation, especially  when  introduced  into  the  duodenum  where  it  may 
neutralize  acid  entering  the  bowel  from  the  stomach  and  thus  spare 
the  liver  and  ])anereas  from  the  effects  of  absorbed  acid.  Underbill 's 
experiments""  with  bicarbonate  feeding  in  diabetes  confirm  these  ob- 
servations. 

One  difference  between  diabetes  melitus  and  phlorhizin  diabetes  is 
that  in  the  former  the  glycosuria  is  due  to  hyperglj-cemia,  the  sugar 
loss  being  an  overflow  like  water  escaping  from  an  overfilled  tank; 
whereas  in  phlorhizin  ])oisoning  there  is  ajiparently  an  hypoglycemia 
— the  loss  resulting  in  this  case,  to  carry  out  the  simile,  from  a  leak  in 
the  bottom  of  the  tank  which  keeps  the  water  at  a  lower  level.  But  the 
results  are  the  same.  ^Moreover,  if  in  diabetes  melitus  we  could  meas- 
ure only  the  chemically  active  or  dissociated  blood  sugar,  it  is  possible 
we  should  again  find  for  this  kind  of  sugar  an  hypoglycemia  compara- 
ble to  that  of  phlorhizin  diabetes.  This  conception  coincides  with  the 
doctrine  that  in  diabetes  melitus  there  is  a  primari)  underconsnmp- 
iion  of  sugar  as  opposed  to  the  idea  of  a  primary  overproduction. 

Overproduction  vs.  Underconsumption. — At  the  present  time  the  chief 
ex])onents  of  overproduction  are  the  followers  of  Kraus,  and  of  von 
Noorden  in  whose  books  ''Die  Zuckerkrankheit"  and  ''New  Aspects 
of  Diabetes"  will  be  found  the  arguments  favoring  this  idea.  A 
translation  of  Minkowski's  criticism  of  the  latter  has  been  made  by 
Lusk."^  In  this  place  it  may  be  briefly  recalled  that  the  chief  argu- 
ments favoring  underconsumption  in  addition  to  what  has  already 
been  said  are  the  following:  (1)  The  respiratory  quotient  in  diabetes 
is  freciuently  found  to  be  low,  and  when  carbohydrate  food  is  admin- 
istered this  quotient  rises  but  little,  wdiereas  in  health  the  administra- 
tion of  carbohydrate  food  results  in  a  greater  rise.''-  (2)  The  acetone 
bodies  (acetone,  aceto-acetic  acid  and  beta-hydroxybutyric  acid)  ap- 
pear in  the  urine  when  for  any  reason  the  quantity  of  sugar  burning  in 
the  body  falls  below  a  certain  minimum,  as  in  starvation,  or  when  a 
})erson  accustomed  to  a  mixed  diet  is  suddenly  switched  to  a  full 
calory  diet  composed  exclusively  of  fat,  or  of  fat  and  carbohydrates, 
with  the  carbohydrate  calories  representing  less  than  10  per 
cent,  and  the  fat  calories  more  than  f)0  per  cent,  of  the  total 
(Zeller  "■'').  In  these  cases  the  restoration  of  sugar  to  the  diet 
abruptly  and  ])ernuinently  stops  the  output  of  actone  bodies.  But 
in  severe  diabetes  the  excretion  of  acetone  bodies  is  less  affected  by  the 

60  Jour.  Amer.  Med.  Assoc,  1017  (OS),  407. 

«i  IMpflieal  Rword.  Feb.  1,  1013.- 

>'-  I'^ir  tlie  literalnre  of  res])irati(>n  sdidies  in  dialtotes  see  Josliji.  Troalment  of 
Diabetes  Melitus,  New  York,  lOlfi;  Du  Bois,  Harvey  Society  Lectures,  1916;  and 
"Studies  from  tlie  Department  of  niysiolo<ry  of  Coniell  University,  lOl.T  ct  scq.; 
|iulilislied  in  tlie  .Areliives  of  Internal  ^Medicine  and  rejjrinted  as  Bulletins. 

«:f  Arcli.   f.   I'liysiol,,   lOI-l,  j).  21:5. 


riiijth'Y  or  i)i\ni:Ti:s  071 

p-ivinj,^  of  siijzar.  l-'ollow  iiii:-  single  l;ii'>ic  doses  there  may  indeed  Ix-  a 
t(Mii])orary  fall  in  the  acidosis,  but  this  is  never  perinaiiently  attain- 
able. One  interpretation  made  of  these  facts  is  as  follows.  In  dia- 
betes there  is  an  acetone  body  output  because  sujiar,  althouuh  brought 
to  the  cells,  fails  to  take  part  in.  certain  cheniical  i-eactions  which  nor- 
mally occur  between  sugars  and  certain  of  the  breakdown  i)roducts  of 
butyric  acid  and  which  normally  prevent  the  diabetic  acidosis.  Hence 
the  bringing  of  more  sugar  has  little  effect.  A)id  iiere  again  one 
might  suggest  that  in  diabetes  glucose  fails  to  interact  with  the  pi-od- 
ucts  mentioned  because  tlie  glucose  is  not  sufficiently  dissociated. 
Another  intei-pretation  has  been  to  the  effect  that  the  sugar  simply 
causes  a  compensatory  decrease  of  the  fat  metabolism,  i.e.,  spare  fat, 
thereby  decreasing  the  formation  of  the  acidosis  bodies.  The  mech- 
anism of  the  process  is  in  any  case  still  a  theme  for  research. 

There  are  numerous  other  theories  of  diabetes,  for  the  presentation 
of  which  the  reader  is  referred  to  the  larger  works.  Lepine  has  long 
stood  for  the  view  that  the  pancreas  secretes  a  glycolytic  oxidizing 
ferment.  Xaunyn's  theory  pays  particular  regard  to  the  ability  of 
the  body  to  "fix"'  glycogen,  while  glycogen  formation  is  held  to  be 
a  necessary  preliminary  step  in  the  utilization  of  sugar.  The  fail- 
ure to  fix  glycogen  he  calls  "  diszoamylie ,"  and  the  other  metabolic 
disturbances  he  regards  as  sequences.  The  complex  develo])meiit  of 
this  same  general  idea  by  von  Noorden,  with  the  added  element  of 
primary  sugar  overproduction,  has  already  been  alluded  to.  Pavy 
saw  in  the  diabetic  a  failure  to  assimilate  sugar;  that  is,  a  failure  of 
the  body  to  incorporate  sugar  in  a  colloidal  combination  which  would 
at  once  permit  of  its  transportation  to  the  points  of  utilization,  and 
prevent  its  prenmture  excretion.  The  assimilation  he  held  occurred 
in  the  villi  of  the  intestines,  and  the  lymphocytes  he  regarded  as  the 
morphologic  elements  which  carry  the  sugar.  Cohnheim's  theory 
that  the  muscles  formed  glycolytic  enzymes,  for  which  the  pancreas 
supplies  an  essential  activator,  is  without  any  substantial  experimen- 
tal support  at  the  present  writing.  Allen  proposed  that  the  ])ancreas 
supplies  an  "amboceptor"  which  is  essential  for  the  proper  colloidal 
blood  sugar  combination. 

Bronzed  diabetes,  the  name  given  to  that  form  of  hemochromatosis 
in  which,  along  with  the  hepatic  cirrhosis,  there  is  an  associated 
fibrosis  of  the  pancreas,  and,  as  a  result  of  this,  the  sym]itoms  of 
pancreatic  diabetes,  will  be  found  discussed  under  the  heading 
"hemochromatosis,"  chapter  xvi. 

Diabetic  coma  is  discussed  under  "acid  intoxication,""  chapter  xviii. 

Lipemia,  which  is  observed  frequently  and  most  severely  in  diabetes, 
is  discussed  in  chapter  xiv. 

THE  END 


INDEX 

XoTE. — Tho  minibcrs  printed  in  bold-face  type  refer  to  pai;es  upon  wliich  the 
topic   is   speeitically   diseiissed. 


AiiDKiuiAr.UKX   roaotioii,    1!»S,    204-207 
specilic-ity   of,   20(j 
witli   veji'etable  proteins,  206 
Abriii,   144,   177,  2-i;5,  225,  293 

poisoning',    liistologie    changes,    146 
Abscess,  4:5."] 

cold  tuberculous,  280 

liver,    135 
Absorption,    337 

impaired,  246 

in  dead  bodies.  338 

partly  due  to  lymph  cliannels,  338 

physical,   240 
Acetanilid,  217.  482 
Acetic,   248,   251,  567 
Aceto-acetic   acid,    550 

poisoning,  553 
Acetone,  550,  567 

bodies,  origin  of,  554 

toxicity   of.   553 
Acetonemia.  558 
Acetonitrile,  592 

test.   604 
Acetonuria,   558,   560,   584 

cachectic,    558 

in   fever,    560 
Acid,  71 

dves,    sul])honic.    50 

fastness.   111.   207 

intoxication.  292.  534,  547-550,  584 
non-di:il)ctic.   557-561 

lactic.  647,  648 

phloretinic,  (i59 

phospliate  retention,   558 

within  cells.  548 
Acidity,  abnormal,  347 

high,  456 

increased,   348 

of  gastric  juice,  245 

of  n\iclei,  46 

relation  of  risror  to.  391 
Acidosis,  74,  412,  549-550,  614 

as  cause  of  death,  560 

at  high  altitudes.  560 

diabetic.  551,  71.  292 

estimation  of.  549 

general.  531 

local.  409 

of  ]iregnancy.  559 

relation  to  diabetic  coma,  552 
Acquired   tolerance.   246 
Acrolein    test,    475 


Acromegaly,   614-616 
Actinosphaerium,    377 
Acute    yellow    atrophy    of    liver. 
379,     403,  '538,     539 
557,  569 
blood  in,  546 
Addison's   disease.   4(i7.   472,   60S 
613-614 
unstriated  muscle  in,  613 
Adenase,  85,  497,  023 
Adenine.  287,  619,  623 
Adenoma,  432 
Adenomatous   goiter,   601 
Adenosine-deaminase.  623 
Adipocere,  399.  410-412 

composition  of.  410 

formation,  411 
Adipose   connective  tissue.  400 

fluids,   302 
Adiposis  dolorosa,  512 
Adiposity,  614 
Adrenals,   445,   60S 

acute  insufficiency.  Oil 

cancers.    503 

choline  in.   123 

cortex,  404 

relation  to  generative  system. 

hypertrophy,  532 

li])oids   in   arterial   disease,   009 
in  ])neumonia.  609 
in  renal  disease.  609 

medulla.  472 

relation  to  carboliydriites.   611 

resemblance     of     ln])erneplirom; 
518 

tumor,  melanotic,  472 
Adrenalin,  609 
Aethalium   septicum.   255 
Agglutination.  183    189,  324 

by  acids.   188 

mechanism    of,    185 

relation   of  salts  to.   186 

relation   to  resistance.   183 
Agirlutinative  thrombi.  325 
Agglutinins.  118,  170,  183-189. 

cell  receptors.  185 

electric   charges   of,    187 

plague.   184 

properties  of.   184 

typhoid.  174.  184 

venom.  154 
Agglutinogen,  183 


100. 
550, 


610, 


608 


I     to. 


.358 


673 


43 


67i 


INDEX 


Aggressins,  130 
Air  einbolisni,  326-327 
Alanine,    495 
AIl)inisui,  4(i7,  577 
A lliiuiKMi- peptone,  258 
Albuminuid   matrix,   437 
Albuminul\  sis,  207 
Albnminous    soaps,    268 
AU.nniins.    21,    351.   350 
Allnuninnria,  349.  417,  563 

alimentary,   528 
Albuminuric  retinitis,  530 
Albumose,  279,  508 

Bence- Jones,  309,  518,  570 
constitution  of.  520 
reaction  of.  519 
Albumosuria,  279,  569-570 
myelopathic,  519  521 
occurrence  of,  520-521 
Alcohol,  50,  105,  244,  257,  301,  483 
cetyl,  514 

efi'ect  of,  on  germicides,  28 
oxidase,  71 
in  liver,  248 
Alcoholism.  412,  595 
acute.  531 
lipemia  in.  413 
Aldehydase,  72,  100,  545 
Alimentary  albuminuria,  528 
levulosuria,  055 

spontaneous,  055 
tract,   247 
Alkali,  71 

albuminate.   258 
dill'usible,  292 
free,  247 

non-difl"usible.  292 
Alkaline  salts,  337 
Alkalinitv,    increased.   440 
of  bile,' 245 
of  blood,  292.  303,   314,  534 

relation  to  bactericidal  power,  292 
total,  301 
real,  291 

relation  of  calcification  to,  439 
Alkaloids,   379 
Abalosis,  598 
Alkaptonuria.    09,    73.    473,    524,    577, 

580 
Allantoin,   357,   023.   025 
Allergy,   193-204 
Alloxan.  020 
Alloxuric    bodies.    019 
Alpha-inicleoproteins.   1 95 
Altmann's  granules.  98 
Aluminium    hydrate,    40 
Amanita   hemolysin,  227 
niuscaria,    140.    147 
jiliallobles.   140.  147.  107 
Amanita  toxin,    147 
Amblyopia.    007 

Ambocei)tor.    211    214.    218.   228 
action  of.  220 
derivation   of,  219 


Amljoceptor.        hemolytic,        219-221, 
232 

properties  of,   219 
immune,  211 

lelation  to  proteins,  214 
stability  of,  213 
union  with  cell,  220 
where   formed,   213 
Amboceptor-complement    bacteriolysins, 

205 
Ambrosia.  147 
Ameb.T,  81,  250 

artificial,  268-271 

taking   of   food,   209 
coli,    135 

relation  of  leucocyte  to,  200 
Ameboid  motion.  39,  254-256 

artificial       imitations      of,      267- 

271 
by  inorganic  substances,  208 
Amibodiastase,  202 

Amino-acids,    20.    279,    281,    311,    546, 
507,  508 
cyclic,  280 

derivatives,  580-581 
nitrogen,  529 
radicals,    176 
Ammonia,  567,  620 
Amnion iacal   decomposition,   455 
Ammonio-magnesium     phosphate.     455, 

450 
Ammonium  carbonate,  251,  520 
compounds,    231 
urate.    258,   454,   455 
Amoeba.     See  Aniebce 
Amygdalin,   280 
Amylase,  07,  SO,  95.  103 

in    urine.    SO 
Amyloid,  417-423,  4(>0 
accumulations,   local,  423 
chemistry  of,  418  420 
en/.yme,  422 
intiUration,    010 
kidneys,  404,  415 

concretions,  423 
origin  of,  421   423 
relation    of   liyaiin    to,   424 
si)lenic,    120 

staining  properties,  420-421 
Amyloidosis.  02,  421 
Anabolic    ))rocesses.    371 
,  Anaerobic      gas-producing      organisms. 
305 
A-iiapbthol.    248 

Anaphylactic    difl'erentiation    of    blood 
corpuscles.   175 
of   hemoglobins.    175 
intoxication.  02.  198 
poison,  character  of,    199 
reactions  with  salvarsan,  109 
shock,    320 
Aiuiphylactin.  202 

relation  of  ]>recipitin  to.  193 
Anaphylactogens,    194 


INDEX 


675 


Anaplivhitoxin,      120,     130,     170,     197, 

by  lUitolysis  of  bacteiia,    1!>8 
formation.    ")(!!• 
from  kaolin  in  lilood,  201 
relation    to   anaphylaxis,    201 
Anapliylaxis,   172,   193   204.  509 
dne  to  protein  cleavaye  \<\   protease, 

202 
pathologic  changes  in,  19!t 
relation  of  anaphylatoxin  to.  201 
Anatomical   and   ciiemieal    fat    changes, 

402 
Ancistrodon  contortrix,   149 

piscivorus,  149 
Anemia,  70,   142,  231.  293,   29S.   371- 
372,  407,  413,  602 
acute,    302 

iiemolytic.  478 
aplastic.  321 

bothriocephahis,    139,  300,  414 
due    to   hemolytic   agencies,    302 
hemolytic,   230 

Ijeruicious,  231,  305-307,  317,  477. 
587 
analysis  of  organs  in,  305 
calorimetric   studies   in,   3116 
cavises   of,   307 
chemical   changes    in,    305 
due  to  hemolytic  poisons,  300 
iron  in  corpuscles,  305 
protein  metabolism  in,  300 
secondary,   225.  294,   317.   322.   300- 

302 
severe.  561 
transitory.   394 
Anemic  heart  murmur,  431 
infarct,  327.  381.  407,  415 
necrosis,  328.   368,  372,  381 
Anesthesia.   558 
chloroform.   78 
Anesthetics.    245 
Aneurism,  322 
Angioneurotic  edema,  351 
Anilin  dve  cancer,  493 
Anilines,"  218.  249,  482 
Animal  parasites,  260.  340,  432 
chemistry  of,  134-143 
eosinophilia    (chemotaxis) .  134 
Anistropic  bodies,  404 

lipoids,    25 
Antagonism  of  ions,  247 
Antelope  dorcas,  463 
Antemortem  rigor,  390 
Anthracene  fractions  of  tar,  493 
Anthracosis,  265 
Anthrax,    276 

bacillus,    108,   131.   132 
Anti-amboceptors,    222 
Anti-anaphylactic    condition,    203 
Antibodies,'  126,  128,  417 
echinococcus,   138 
effect  of  light  on,  374 


Antibodies,    hemolytic,    in    vitro    tissue 
cultures,   170 

relation  of  lipoids  to,    169 

specific,    therapeutic    stimulation    of, 
174 
Anticatalase,   70 
Anticoagulin,    318 
Anticomplement,  239 
Anti-endotoxins,    130 
Anti-enzymes,        63   68,        89,        277, 
289 

specificity   of,   66 
Antiferments,  291 
Antigelatinase,   117 
Antigen-antibody-com|)lement,  202 
Antigenic  activity,   171 
Antigens,  166-171 

etl'ect  on  non-stiiated  muscle,  201 

excess  of,   190 

from  echinococcus  lijjoids,  168 

from   ta])e   worm    lipoids,    168 

from  tubercle  bacillus,   168 

immunological    specificity   of,    173 

in  vegetable  proteins,  204 

monovalent,   179 

nature  of,    166 

non-specific.    235 

several  in  single  organism,  174 

specific.  235 
Antihemolysins,  222,  231 
Antiketogenesis,   554 
Antiketogenic  agents,  554 
Antikinase,  64,  319 
Antilijiase,  thermostable,  79 
Antimony,   247 
Antioxidase,  467 
Antipepsin,   65 
Antiplatelet  serum,  239,  300 
Antiiuioumin.   69 
Antiprotease.  361 

in  bacteria,  118 

serum.  277 
Antiy>^■retic  drugs.  259 
Antipyrin.    160,   243,   258,   560 
Antirennin.  64 
Antisensiti/ation.   203 
Antiseptic  substances.  378 
Antiserum,  171 

cholera.  211 

excess  o^,  190 
Antithrombin,  295.  308,  316 
Antithrombin  -  prothrombin       balance, 

321 
Antitoxin.  177-183 

difl'usion  of.   182 

filterability    of.    182 

against  cantharidin.  170 

associated  -with  pseudoirlobulin.  181 

chemical   nature  of,   180 

diphtheria.   174 

Ehrlich's  theory  of.  177 

for  endotoxin.   130 

neutralization  of  toxins  by.  179 

putrefaction  of,   182 


676 


INDEX 


Antitoxin,     relation     of,     to     enzymes, 
180 
to  proteins,   181 
specilicity   of,   ITS 
Antitrypsin,  63-  68 

content  of  blood,  200 
Antitryptic   titer,    535 
Antivenin,   155,   156-157 
Ants,  160 
Anuria,  344 
Aorta,    atheromatous    degeneration    of, 

012 
Aphrodite  aeuleata,  170 
Aplastic  anemia,  321 
Aporrhegma,   122 
Appendicitis,  433 

chronic,  414 
Arabinose,   649 
Arachidic  acid,  514 
Arachnolysin,  159 
Arcns  senilis,  415 

Arginine,  20,  84,  90,  100,  285,  544,  545, 
581 
nitrate,  544 
Aromatic  compounds,  chromogenic,  613 

radicals,  176,  194 
Arsenic,  165,  244,  246,  542 
fixation  to  nucleus,  246 
immunization   against,   247 
poisoning,  342,  539 
sulphide,   34.  41 
Arseniuretted  hydrogen,  217,  232,  485 
Arterial  degeneration  from  epinephrin, 
612 
disease,  adrenal   lipoids  in,  609 
Arteries,  large,  439 
Arteriosclerosis,  416,  424,  428,  584,  587, 

610,  612,  614 
Arthritis,    357 
gonorrheal,  433 
rheumatoid,   629 
Arthroi)athv.   tabetic,   351 
Artificial  a'mel)a',  268-271 

takinir  of  food.  269 
Ascaris,  140-141 
chemistry  of,   141 
luml)ricoides,    135 
megalocepliala,    140 
pioducts  of  metabolism,   141 
toxicity  of.   141 
Ascites  adii)osus,  364 
chvhnis.   340 
lluid.  311 
liemorrhagic,    490 
Aseptic    li<|uefaction    necrosis,   276 
Asiatic   cliolera.   560 
Asparaginic  acid.  495 
Aspartic   acid.    544.    584 
Asphyxia,  263,  390.  538 
Asphyxia!    conditions,  560 
Asphyxiation,     346 
As|)irin,   169 

Astlienia.   gastro-int(>stinal.    lil  1 
muscular,   614 


Asthenic    uremia,    529,    532 
Asthmatic  sputum,  311 
Atheroma.  414 
Atheromatous  areas,  441 

degeneration  of  aorta,  612 

masses,  cholesterol  in,  25 

patches,  405,  415,  459 
Athyreosis,  592,  598 
Atophan,  630 
-^toxvl,    169 
Atrophy,  92.  393   394 

acute  yellow,  100.  379,  403,  538,  539- 
550.   557,  569 

brown,  393,  474,  482 

of  kidneys,  474 

of  liver,  474 

of  pancreas,  393 

simple,  393 

a^-ray,  of  ovaries,  376 
of  testicles,  376 
Atrojjine,   560 
Autocvtotoxins.  240 
Autod'igestiun,   82,  318 
Autohemolysin.    233 
Autoimmunization.  311 
Autointoxication,  523-565 

gastro-intestinal.  566-589 
Autolysis.  82-105.  317.  327,  368 

bacterial,   rate  of,    119 

defense  of  cells  against,  88 

histology  and  chemistry  of.  99-100 

inlluence    of    cliemicals    in,    86 

liver,  efi'cct  of  thyroid  on,  591 

of  ])acteria.  118-120 

of  cell  proteins,  403 

of  leucocytes.  308 

of  tumors,  496 

pancreatic.  386 

products  of.  279 

relation  of  age  to.  88 
to  asi)hvxiation.  89 
to  toxins.  102-103 

witliin  Ixnly.  370 
Autolytic   enzymes,   67 
of  tissue  cells,  277 

products,  bactericidal  ]>ower  of,  119 

ju'oteolysis.  526 
Auto-o]isonin.  233 
Auxanograi)hic    nu^thod.    114.   502 
Auxetics.  493 

Bactt.t.is.  acid-fast.  207 

aerogones  capsulatus.  307.  327,  389 
anllirax,    108,    131,    132 
botiilinns.  177.  585 

toxin.   16S 
cholera'  gallinariuni.  257 
coli   communis.  70.  94.   132.  264,  289, 

320.  3S9.  451.  574.  633 
diphtheria.    70,     177.    131,    132.    259, 

320 

lecithin    in.   Ill 
eTni)h\senn»1osns.  478.  482 
enteritidis.   122 


INDEX 


677 


Bacillus,  iMicdlaiuU'r's,  258 
(iiirtuiT,    17:5 
liu^'-cliolora,    .■524 
niegatliorium,   22.') 
l>aratyiilioicl,    17:5 
plajiue,    107 
prodi-iiosus.    114,   320 
]iroti'Us.    Ill) 

Uiiorosceiis.     l."{2 
l.vo.'vanevis.    11-1.    132,    177.   224.   259, 

27S,  320 
siihlilis,    114 
tetanus,  70.  177 

tubercle,    70,    70,    00,    107,    131,    132, 
204.  320 
antijien   from,    108 
foiiipdsitiou   of.    lOS 
fats  of.  110    111 
fatty  acids  in,   112 
modification    of    acid    fastness    of, 

111 
no  cholesterol  in.   111 
phosphatids   in,    111 
typhosus.  70.  120.  132,  183.  211,  214, 

224,  2G4,  320,  451 
xerosis,  320 
Ba  CL,  200 
Bacteria.    73,    411 
anti-protease  in.   118 
autolysis  of.  89.  90.  118-120 
chemical  comjiosition.    107-113 
chemistry  of.   106-113 
chemotaxis  in.   107 
cholesterol    in.    Ill 
chromatin   of.    100 
com])osition  of  cell  wall,  110 
decolorization  of.  112 
ectoplasm  of,   106 
endoplasm  of.   106 
inhibition  of.   102 
Gram's    method    of    staininfr.     112- 

113 
motile.    250 
nucleus  of.   100 

patlio.acnic.   endotoxins   in.    119 
plasmolysis   in,   107 
plasmophysis  in.   107 
putrefactive.   506 
reducinji   power.   114 
relation  of  nutriments  to.  lOS 
staining  reactions.   112-113 
synthetic  activity  of,  100 
tolerance  of.  252 

typhoid  colon.   difTerentiated  by  acid 
agglutination.   188 
Bacterial  autolysis,  rate  of.  110 
carbohydrates.  109-110 
caseins.  108 
catalase,  117 
cell      wall,      animal      or      vegetable, 

110 
cultures,  sterilized,  258 
decomposition.    10 
digestion,  products  of,  116 


Bacterial  endotoxins,  118 

enzymes,    113    120,   27i;,   3S2 
immunity  again^l,   117 
relation    to    pathngonicity,    110 
role     of,    in      infectious     diseases, 

110 
fats,  110   112 
and  fatty  acids,   111    112 
ell'ect  of  media  on,    1 1 1 
formation,  390 
nucleic  acid,    109 
nucleoprotcin,    108 
pigments,  132-133,  280 
alcohol  s.>inl)lc,  133 
insoluble,    133 
Avater  soluble,   133 
])oisons,    71,    502 
])recipitins,   192 
proteins.   270,  280 

poisonous.    131-132 
toxic  not  specilic,  132 
toxicity  of,  131 
proteolytic    enzymes,    resemblance    to 
trypsin,  110 
power,  relation  to  virulence,  117 
spore,  367 
substances,    195 
toxalbumins,    108 
toxins,  125,  165,  265,  318,  407 
Bacteriolysins,    118.    128.    358 
amboceptor-complement,  265 
Bacteriolysis.  207,  213.  214 

serum,"  208,  210-214 
Bacteriolytic    endolysins,    280 
Bacterium   ternio,   254 
Barium,    240 
Barlow's  disease,  446 
Basal  metabolism,   590,  602 
Bee  poison.    160 

toxolecithidin   in.    100 
Bence-Jones   albumose.    300,   518,   570 
constitution   of   520 
reaction  of,  519 
Benzene.  317 
Benzoic  acid,  250 
Benzol,  259 
Benzopurpurin,  493 
Beriberi,  286 

Beta-iminazolyl-ethylamine,   576,   014 
Beta-oxvbutric  acid,  550 
Betaine."  123,   124 
Bezoar    stones.    463 
Bibasic  urate.  620 
Bichloride  of  mercury,  43 

poisoning,  532 
Bile,  127.  232 

acids.  218,  246.  251 

alkalinity   of.   245 

colloids,  "electro-negative,  452 

fat  content.  326.  403 

pigments.   229.  484.    486-487.   560 

calcium   salts  of.  448 
salts,   487.  506 
thickening  of,  453 


678 


INDEX 


Bile  thronil)i.  485 
toxicity  of.   486 
tracts,  inflammation  of,  453 
Bile-chicts.  casts  of,  450 
Biliary  calculi.  448-454 
cirrhosis,  486 
fistula.   558 
Bilicyanin,  450,  484 
Bilifiiscin,    449,    484 
Bilihumin,  449,  450,  484 
Biliprasin,  484 
Bilirubin,     296,     375,     449,     474,     477, 

484 
Bilirubin-calcium   calculi,  mixed,  449 

])ure.  449 
Biliverdin,  375,  449,  484 
Biliverdincalcium,  449 
Binding    substance,    454 
Biurate,  620 
Black  flies,  160 
Blackwater  fever,  232 
Bladder.  457 

urinary,    423 
Blastomycetic  dermatitis,   274 
Blisters,   burn.   565 

fluid.   362 
Blood.  247 

alkalinity   of,    292,   303.    314.    534 
relation  to  bactericidal  power,  292 
total,    301 
amount  of  water  in.  331 
antitrypsin   content,   200 
buffer  value,  291 
cadaver,    320 
calcium  in,  440 
cells,  red.     See  Erythrocj/tes 

white.     See  Lencori/tes 
changes    after    hemorrhage.    294 

in  passive  hyperemia.  313 
chemistrv   of.   in    leukemia.    308 
coagulability  of.  200,   316.  321-322 
in  disease.  319 
modification.   317  318 
retarding  of,  318 
composition  of,  289 
corjuiscles.    aiiaphviactic    differentia- 
tion of.  175 
red,     70,     289.     See     also     Eryth- 

rnrj/tes 
white.     See    Levrofytefi 
effect  of  venoms  on.   153 
electrolyte  concentration  of.   290 
enzvmos.  291 
extravasatod,   29.-)  297 
fat  in,  326 

fibrin  content,  in  dispiisc.  321 
formed,  elements  of.  289.  290 
freezing-point.   301.   308 

lowering  of.  5f)6 
imbibition  of,  412 
in    acute   Acliow   atrophv.   546 
hiking  of.'  215 

leukemic,  coagulat  inn   of,  308 
lipase,  535 


Blood,  menstrual.  320 

nitrogen,  non-jjroteiii,  534 
pigments,  476  484 
plasma.  290 

increased  filterabilitv  of,   344 
reaction   of.    291-292 
relation  of  hinjjli   to.  ."i.'JO 
platelets,  289,  290,  29!),  ;517 

poisons.  31S 
))oisons,  406 
pressure.  298,  331,  349 
high.  612 

increased.   341-342 
intracapillary.  334 
]u-oteins,  290 
serum.  200 

normal.  486 
state  of  sugar  in.  643 
sugar.  641 

concentration,  641 
content,  535 
toad,   ]>nison  in.   161 
viscosity  of.   292-293 
Bloody  diarrhea,  563 
Body  temperature,  subnormal.  615 
Boiling-point,   elevation   of,   of  colloids, 

38 
Bone-marrow,  91,  296,  311 

tumors.  570 
Bones,  congenital  fragility,  447 
Botliriocephalus      anemia,      139,      306, 

414 
Botulism.  122,  567 
Brain.   77,   83 
edema  of.  535 
enzymes  in.  78 
softening,  342 
tissue.    123 
tumor  of,  531 
wet,  531 
Breast,  cancer  of,  437 
Brick-dust   deposit,   455 
Brilliant       green,       sensitization       by, 

220 
Bronchi.  423 
Bronchial  catarrh,  434 

secretions.  404 
Bronchiectasis,  281 
Bronchiectatic  sputum.  414 
Bronchitis.  281,  450-454 

pufrid.  434 
Bronzed  diabetes,  483.  671 
Brown  atrophy.  393,  474,  482 
Brucin.  251 
Bruise.  295 
Bufagin.  161 

Buffer  value  of  blood.  291 
Bufo  agua.   161 
Bufonin,   161 
Bufotalin.    161 
Burn   blisters.   565 
Burns.    S5 

hemolysis  in.  232 
leucocytosis  in,  261 


INDEX 


07!) 


Burns,  siiiifiliiinl.  imisoiis  produced  in, 

562   565 
Butter,  2S7 
cysts,  r.l.") 
Butyl    chloral.    -J  1!» 
Butyrase,  77.  7'.t 
Butyric  acid,  ^u,  M7 

Caciiixtic    acetoiuiria.    008 
conditions,  2!)4,  .S;)4 
diseases,  oOG 
edema,  ."$44 
Cachexia,  (Hi,  711,  2SI,  302,  417,  .")(iO 

cancer.  :5()!».   492 
Cadaver  blood,  320 
Cadaverine.  123,  5*57,  582,  583 
Caffein,  248,  (il9.  (327 

demetliylated  in  liver,  247 
Caisson  disease.  32(5 
Calcareous  stones,  450 
Calcification,     325,      329,     424,     435- 
447 
chemistry  of.  439  441 
deposits,  coniixisitioii  of,  436-437 
metastatic,  438  439 
of  gan,u;lion-cells.  43S 
of  renal  epithelium.  438 
patholo-rical.   415,   435,   438-439 
pt'ricai'dial.  43(1 
]ihi's|)lioric  acid   in,  442 
relation  to  alkalinity.  439 
to  ossification,  435-436 
to  retrogressive  clianues,  440 
Calcified  areas,  structure  of,  437 
]ilaqiu's,  323 
tidiercdes,   464 
Calci'^ving  lii)oma.  441 
Calcium.   251.   285.   303,    308.   456,   487. 
499 
carhoiuite  calculi,  458 
chloride,  336 
oout.  439 
in  blood.  440 
in   fibrin  formation.  316 
oxalate.  454.  45G.  457 
calculi.   457 
L'allstoiies.  450 
salts.  48.  317 

absorption  of.  442-443 
effect  of.  on  phajjocytosis.  263 
of  bile  pigments.  448 
soaps.  387 

formation  of.  441-442 
Calcium  iron  incrustations.  437 
Calcospherites.  437 
Calculus  be/oar.   463 
biliary,  448-454 
cali'areous,  450 
calcium  carbonate.  458 

oxalate.  457 
cholesterol.  459 

crystalline.  452 
cystine.   458 
fecal.  463 


Calculus,  librin,  459 
fusible,  458 
in  Chiiui.  456 

iiulifjo,  458 

intestinal,  462  463 

luni;-.  459,  4(i4 

metaniorpliosed,  455 

mixed    bilirubin-calcium,   449 

oxalic,  584 

jiaiHTeatic,  461   462 

l)hosplialc.  457  458 

pure  liilii'uhin-calciuin.    \  19 

salivary.  462 

str\ivi(.  457 

urate,  456-457 

uric  acid.  455   456.  027 

urinary,  454  460 

disinteoration  of.  459-460 
general   jfroperties,  459-460 

urostealith.  458 

xanthine,  458 
Calories,  281 
Calorimetric  stiulies   in   ])eriii(i(ius  aiu»- 

mia,  306 
Calves'  brains,  585 
Camphor.  249 

Cancer.  66.  71,  79,  94.  230.  231.  292, 
322,  344.  356,  424,  425.  432, 
497,  560.  569.  634 

adrenal.  503 

anilin  dve.  493 

cachexia.  309,  492 

chemical  stimuli  causing.  493 

colloid,  427,  516 

extracts,   104 

hemolytic,  492 

glycyl-tryptoidiane  test  for.  501 

hemolysis  in,  504-505 

increase  of  Oil  in  blood  in.  507 

induced       cell       reproduction       and, 
493 

lysins,  241 

metabolism  in,  505-507 

of  breast.  437 

o*^  stomach.  104.  504 

of  thyroid.  604 

putrid.  574 

skin,  in  colored  races.  467 

squamous  cell,  516 

a"-ray,  377 
Cantharides.  312 

blisters.  260 
Cantharidin.  340 

antiioxin  a>jainst.  170 
Caoutchouc  colloid.  426 
Capillaries,  injury  to.  254 

obsl  ruction.  326 

permeability  of,  333-334.  349 

walls.      increased      permeability      of, 
342   344 
Capra  aecraurrus.  463 
Carbohydrate    uu-talxdism.    602.    635- 
671 
effect  of  tlnroid  on.  591 


680 


INDEX 


Carboliydratos.  :V.],  35,  25S,  335,  551 

aiitiketouciiic  act  ion.  37'J 

bacttM-ial.   109    110 

colloidal,   34(j 

fermentation    of,    products   of,    583- 
584 

in  food,  fat  from,  398 

relation  of  adrenals  to,  611 

tolerance,  increased,  615 
Carbolic  acid  gangrene,  378 
Carbon  dioxide,  53,  293,  326,  501.  5Q7 
Carcinoma.     See  Cancer 
Carcinomatous  exudate,  304 

peritonitis.  ;553 
Cardiac  defects,  5(il 

disease,  71 

with  edema.  293 

dropsy.  345,  349 

edenui,  348 

failure,  acute,  610 

incompetence.  324,  350 
edema  o*',  341 
Cardiolvsin,  241 
Caries. '438 
Carotid  gland,  617 
Carotin.  448,  475 
Cartilase.  normal.  420 
Caseation.  98.  99.  382-384,  415 
Casein.   195,  282 

bacterial.  108 
Caseous  areas.  438,  441 
encapsulated,  415 

material,  fat  in,  383 

tubercles,  276 
Castration,  444 
Catabolic.     See  KatahoUc 
Catalase,    60,    67,    69-71,    280,    502, 
595 

bacterial,  117 

in  urine,  71 
Catalytic  agents,  56 
Cataphoresis.  38 
Cataract  diabetic,  375 
Catarrh,  bronchial,  434 
Catarrhal  inflammation,  427 
Cations,  28 
Cell,  acids  within.  548 

autolysis,   inti'avitam.  408 

chemistry  and  i)]iysics,   17-52 

colloid   eipiilibrium   in,   373 

dead.  eflVct  of  dyes  on.  370 

death,  43 

from  d'CTuicals.  378 
physical  changes  in.  370 
recognition   of,  307 

divisifni.  23 

imitation   of.  271 

endothelial,  219 
death  of.  343 
lar<re,  273 
sudanonhile  change,  272 

epithelioid.  273 

foamv  phatrocvtic,  405 

functioning,  367 


Cell,  giant,  273,  416,  631 

giant,  273,  416,  631 
formation  of,  273-274 

globulins,  coagulation  of.  373 

inorganic,  constituents  of,  26 

Kupfler,  474 

nerve,        coagulating        temperature, 
372 

nutrition   impaired.  497 

pancreas,  emboli  o'',  387 

piiagocytic.  219 

physical  chemistry,  26-43 

j)roteins  of.  21 
autolysis  of,  403 

re]iro(luction.  367 

induced,   cancer  and,  493 

sap,  30 

sex,  174 

state  of  suoar  in,  044 

structure  of.  43-52.  338 

tissue,  autolytic  (Mizymes  of,  277 
iK'havior  of,  273' 

tumor,  404 

wall,  IS.  30.  49   51 
Cellular  immunity.  244 
Cellulose,  30.   109 
Centipedes.    159-160 
Central  necrotic  areas,  273 

softening.  325 
Centrosomes.  47 
Cephalin.  24,  316 
Cephalonods.  576 
Cerebral  edema.  532 

localized.  531 
Cerebrin.  279 
Cerebrospinal  fluid.  65 
freezina  noint.  360 
Cestodes.    137-140 
Cetvl  alcohol.  514 
Chalicosis.  405 

Charcot's   crystals,   311-312 
Cheese,  3S2  " 

ripening  of,   399 
Cliemical  alteration.  240 

combination.  240 

lUMitralization  by.  523 

composition  of  ])rotcin  molecule.  20 

defenses.  245 

react i<ms,  2^> 

stimulus.  284 

causing  cancers,  493 

substances,  abnormal  toxic.  523 

transformation.  523 
Chemorecentors.  243 
Chemotactic    substances,    non-hacterial, 

257   259 
Chemotnxis.   62.   254   261.   314 

necatiye.  257 

o*"  bacteria.  107 

of  leucocytes,  250 

nositive.  250    271,  270 

theories  of.  266-275 
Chomot  ronism     255 
Chimi>anzee,  625 


INDEX 


681 


Chitiii,  ;i(),   110,  i;Ji 
Chitosamia,  513 
Chloial,  258 
Chlorides,  335 

retention  of,  345.  350 
Chluroforni,  5(i,  257 
anesthesia.   78 
narcosis,  540  541 
necrosis,   4iis 

of  liver,  320 
poisoning,  37'J,  53!),  557 
Chloroma,   47t) 
Chlorophyll.    4S(I 

Chlorosis,  302   304,  344,  587,  G12 
etiology  of,  303 
iron  in,  304 

starvation  canse  of,  304 
Cliolagiigue  action,  488 
Cholalic  acid,  463 
Cholelithiasis.  453 
Cholemia,  488,  490,  567 
Clioleprasin,  484 
Cholera,  11!) 
antiserum.  211 
Asiatic,  560 
vibrios,  70,   120 
Cholesteatoma,  515 
Cholesteatomatous  tumors,  415 
Cholesterase,  79 

Cholesterol.    23-25,    85.    100,    138,    217 
226,  236,  237,  244.  290,  297 
303,  357,  361,  401,  404,  410 
413.  448,  545,  609 
calculi,   459 

crystalline,  452 
laminated.  449 
pure,  449 
esters,   25,   415 
gravel,  450 
in  bacteria,  111 

pathological  occurrence,  415-417 
relation  of  ]ihagocytosis  to.  263 
steatosis,  405 
tests  for.  415-416 
Cholesteroleni ia .  4 1  (I — 1 1 7 
Choline,    93.    123-125.    360,   501,    562, 
567,  584,  609 
in  adrenals,   123 
Choluria.  490 
Chondrin.  34,  417 
T'liondrodystropliia  f(ptalis.  603 
Cliondr()itin-suli)liuric  acid.  418 
Chondroma,  432,  511 
Chondrosin,  513 
Chorioepithelioma,  432,  516 
Chromatin,  44 
of  bacteria,  106 
threads,  47.  48 
Chromatolysis,   562 

of  cortical  ganglion  cells,  531 
Chromium  ]>oisoning.  477 
Hiromogen.  white.  469 
Chromogenic       aromatic       compounds, 
613 


Chyle.  lonipo.Nilion  ol',  362 
lat  from,  412 
relation  to  lymph,  331 
Chyliform  lluids,  302 
Chylothorax,  340 

fluid,  363 
Chylous  ascites,  340 

clVusions,  362-364 
Chyluria.   ."Uit,  36;i 
Ciaccoi's  method,  402,  405 
Cilia,  275 
Ciliated  ei)itlielium,  238 

proto/oa,   255-256 
Circulation,   338 

feeble,  440 
(  ircuhitory  disturijances,  595 
Cirrhosis.  78,  92,  356,  587 
biliary,  4S6 
of  liver,  320,  355 
Citric  acid,  251 
Cloudy  swelling.  394-396 
CO^  content,  decrease  in,  440 
Co-agglutination,    186 
Coagulated  protein,  383 
CoagT-ilating  temperature  of  nerve  cells, 

372 
Coag-ulation  of  blood.  200,  298,  302,  316 
decreased,  546 
in  disease,  319 
modification  of,  317-318 
retarding  of,  ."ilS 
necrosis.  381-382 
of  cell  Liloliulins,  ;i73 
of  coll.iids.  40 
of  leukemic  blood,   308 
reaction,  238 

time  of  blood.  219.  321-322 
Coagulins,  316,  317 

tissue.  323 
Coal-jngment.  465 
Cobra    poisoning.    152 

venom,    G>^,    102,    150.    151,    227,    228, 
241 
resistance,  505 
Coccus.   Cram-positive.   207 

infections.  422 
Cod  liver  oil.  287 
Coelenteratts,  poisons  in.   164 
Ccenurus   cerehralis.    138 

scrialis,  138 
Co-enzymes.  61 
Cohesion  allinity,  267 

])ressure,  2(i7 
Cold,  effect  on  life,  373-374 
tuberculous  abscesses.  280 
Cold-blooded   vertebrates,   195 
Collagen.  424 

Colloid.  32.  34-43,  318.  334,  448 
altered  hydratioTi.  254 
anu)rj)hous  form.  35 
bile,   electro-negative.  452 
cancer,  427,  oK! 
caoutcliouc,  426 
capacity  for  water,   336-337 


682 


INDEX 


Colloid  dianges  with  time.,  371 

cliaii^es  with  time,  371 

cliarafteristics  of.  35   39 

(U'^cncratioii,   425-428 

depression    of    freeziugpoiiit,    38 

diffusion  of,  38 

effect     of,     on     chemical     processes, 
3!) 

electrical  phciioineiia  of,  38 

elevation  of  lioiling-point,   38 

emulsion,  452 

goiter,  iodin  content,  600-(301 

liydrophilic,  330,  409 
tendency,   343 

molecules,   395 

non-dillusihility  of.  37-38 

osmotic  pressure,  38 

ovarian,  513 

permeability  to,   371 

precipitation,   187,   301 
and  coaj^ulation,  40,  41 

serum,  electro-positive,  452 

soluhilitv  of,  36 

structure  <if.  42.   43 

thyroid,  426,  593 

tissue,    increased    livdration    capacitv 
of,  346   347 
Colloidal  absorption,  349 

carbohydrates,    346 

hydration,  347 

material,   454 

nitrogen,   506 

poisons  in  tumors,  503 

suspensions,  40 
Colon  bacillus.  70,  289,  320,  451,  574 
Colostrum,  220 
Colubridae,   148 

venenosa",   148 
Coma,  487 

diabetic,  412,  550-554,  558 
relation  of  acidosis  to,  552 

uremic.  355 
Combination.  248 
Complement,    210,    211-214,    218.   358 

deviation,  234.  507 

fixation,    190,   234   238 

hemolytic,   21!).   221-222 

indicator.  234 

origin  of,  21 1 

reaction  to  chemicals,  212 

relation  to  enzymes,  211 
Complementoid.  212 
Concentric    hiiniiiafed    structure,    448 
Concretions.  447  466 

cutaneous,  465 

cystine,   582 

nucleus.   447 

prejmtial,   463-464 

prostatic.  464 

tonsillar,  405 

urinary,     general     ]irn|icrties.     459- 
460 
Conductivity.      relation       to      freezing- 
point.   290 


Congenital   fragility   of   bones,  447 

hemolytic  jaundice,  231 
Congestion,   passive,   314,   333 

of   liver,  431 
Conglomerate  stones,  450 
Conglutination,    186 
Conglutinin  effect,  223 
Congo-red,   493 
Connective-tissue    h\alin,    423-424 

mucin.  428 
Constiiiatioii.   570 
Convulsions,   531,  538 

epileptic,  562 
Convulsive  poisons,   87 
Copepods,  256 
Copper,   223 

nxation,   246 

metallic,  258 

sulphate,  29 
Copperhead   venom,   228 
Coral  snake,  148 

Corpora  amylacea,  423,  425,  452,  460- 
461 
of   lateral   ventricles,   461 

fibrosa,   424 
Corpus  luteum,  404,  474,  475,  608 

scars,   480 
Corpuscles,   pus,   278 
Crab  poison,  164 
Crayfish,  extract  of,  332 
Creatin,  85,  92,  561,  614,  616 
Creatinin,  525,  529,  561,  627 
Cresols,  248,  567,  576 
Cretinism,  586.  590,  601-604 
Crotalus,    149 

v'enom.  151 
Crotin,  144,  177.  223,  225,  276,  293 
Crystalloid,   27-34.   448 

lymphagcgues.   332 
Crystals,  "Charcot's,    311-312 

margaric   acid.   414 

margarin,   389,   414 

spermin,  311 
Cupping,  342 
Curare,  245 
Curcin,    144.    145.   225 
Cutaneous  concretions,  465 

epithelioma.   516 
Cyanosis,   550 

enterogenous.   580 
Cyclamin.  227 
Cyclic  amino-acids,  2S6 

compounds,    370 

ketone.    530 

vomiting.   557,  559   560 
Cylindroma.   425 
Cvstadenoma.   ovarian.   51:5 
Cysteine.  251 
Cystic  goiter.  601 

"thyroid.  426 
Cysticercus   ])isiformis.    138 

tennicollis.    138 
Cystine.  20,   581    583 

calculi.  458 


INDEX 


6813 


Cystine  cdiicrctiuns,  jS'i 

sulphur,  oxidutiun  of.  (il4 
Cystiiiuria,  524,  577,  582  583 
Cystitis,   457 

Cystoma,  ])roliferating,  512 
Cysts,   butter,   515 

dermoid,   415 
of  ovary.  514 

licMiorrliajiic,  42G 

liydatid,  432 

iiitralijj;ameiitary    papillary.    514 

ovarian.  I'ontents  of,  512 
muiH)ids  of.  513 

])arovarian,   514 

serous.  512 

soap,  515 

tubo-ovariaii,   514 
Cvtolvtic  action,  488 
Cytoplasm,  18.  46-49 

structure  of,  46-48 
Cytosine,  019 
Cvtotoxic    influence.   508 
Cytotoxins,    214-215,    358 
Cvtozvme.   310 


Dakin's  racemized  protein,  106 
Deaminization.  116 
Deaminizing    enzymes,   501 

power.  014 
Decalcification,  443 
Deculorization   ot"  bacteria.   112 
Decomposition,  bacterial,   19 

of  fats,  products  of.  584-585 

tension,  380 
Defensive   ferments,    205 
Deficiency  diseases,   287 
Degeneration,    arterial,    from    epineph- 
rin,  612 

atheromatous,  of  aorta.  012 

colloid.  425-428 

fatty.   79.   3!»7 

diic  to  i)()isons.  407 

liyaliiu'.   :(S2.   423-425 

lardaceiuis,  417 

lijx.idal.  410 

liver.    100-102 

mucoid.  420.  427  428 

of  elastic  tissue.  371 

))arencliymatous.    395 

reaction  of.   392 

red,  498 

waxv,  379.   392  393 
Dehydration.   251.  429 
Delirium,   503 

tremens.  405.  531,  009 
Dementia  priecox,  230,  575 
D'-rmatitis,   586 

hlastomycetic.   274 
Dermoid  cysts,  415 
of  ovary.   514 
Desoleolecithin,  228 
Deutero-alhumose,  309,  544 
Deutero-proteose,  543 


Dextrin,  34 

urinary,  388 
Dextrose,  429 

Diabetes,  09,  74,  7.S,  2!I3,  322.  300,  412, 
410,  417,  002,  007,  t)12,  010, 
03.1-071 
bronzed,  483,  071 
glycogen   iiililtration   in.  434 
insipidus.  ()15 
mellitus,   665   671 

overproduction     I's    underconsump- 
tion   in,    070 
t)ther  theories,  071 
paiH-rcatic,   412.   665-671 
phlorhizin.  493,  557,   659   664 

metabolic   phenomena,    0(i2 
theory.  (iOS 
Diabetic  acidosis,  71,  292.  551 
cataract,   375 

coma,  412,  550-554,  558 
relation  of  acidosis  to,  552 
Diamino-acids,  21 
Diamino  nitrogen,   545 
Diarrhea.   245,   584 

bloody,  563 
Diarrheal  disorders,  570 
Diastase,  61,  07,  291,  362 
Diasthesis,  uric-acid,  627 
Dibothriocejjhalus   latus,    139 
Dielectric  constant  of  water,  38 
Diet,  effect  o*'  tumor  growth,  493 
Difflugia    shell,   270 
Ditlusion.  29-33.  337 
Digestion.   171 

bacterial,   products   of,    110 
normal,   products  of,   568-570 
Di<festive   ferments.  506 

iluids,  567 
Digitonin,    220 

Di-iodo-di -hydroxy-indole,   595 
Diose,  645" 
Dioxv])henols,  507 
Diphtheria,   382 
antitoxin.   174 

bacillus.     70.     131.     132.     177,     259, 
320 
lecithin  in.   Ill 
toxin,  75,  87,  102.  103,  127,  128.  218, 
200.  317 
digestion  of.  127 
toxin  antitoxin.  70 
Disassimilatorv  hormone,  593 
Dispersion  of  leucocytes.  273 
Dissociated  jatindice.  489 
Distillation   of  wood.  493 
Distilled  water.  257 
Diszoann  lie.   071 
Ditlrich's  plugs,  389 
Diuresis,  relation  of  ions  to,  28 
Diuretic   substance,   614 
Diver's    palsy,    326 
Dilactic  acid,  94,  280 
Doubly  refractive  lipoids,  25 
Dracunculus,    143 


684 


INDEX 


IJiopsv,  345 

cardiac.   :i45,,   349 

renal,  344 

soda,  350 
Drug  poisoning,  relation  of  lactic  acid 
to,   55(j 

tolerance,  244 
Drug-protein   compounds,   243 
Ductless  glands,  447,  616-617 
Dyes,  absorption  of,  338 

basic,  increase  in  aliinity  for,  285 

ell'ect  of  on  dead  cells,  370 

fat   solubility,  50 

fatty  acid,  414 

organic,  34 

precipitins  fur,    170 

sulpiionic  acid,  50 
Dysentery  toxin,  127 
Dystrophy,   muscular,   393 


EcHiNOCoccus  antibodies,  138 

cvst  fluid,  chemistry  of,  138 
'  wall,   138 

fluid,  complement  fixation,  138 

lipoids,  antigen  from,   168 

polymorphus,    138 
Eck's  fistula,  52G 

Eclampsia,   70,   321,   379,  533-539,  569, 
586 

chemical  changes  in.  533 

etiology  of,   535 

puerperal,  78,  539 

relation  to  uremia,  533 
Ectoplasm  of  bacteria,   106 
Ectosarc.  266 
Edema,   314,   330  366,   382,   395,   531 

acute.  340 

angioneurotic,   351 

cachectic,  344 

cardiac,  34S 

causes  of,  339  352 

cerebral,  532 

chronic,  340 

ex  vacuo,  342,  382 

lluids,   physical  chemistry,  354 
])rotcin  contents,  356-357 
reaction  of,  356 
toxicity  of,  357-358 
varieties  of,  359-365 

in  luMiis,  341 

inlhmmiatorv,  253,  340,  343,  350-351 

intercellular,  337 

intracellular,   337 

localized   cerebral,  531 

neuropathic,   351 

neurotic,  345 

of  brain,  535 

of   cardiac   incompetence,   341 

of  liver,  340 

of   iieplircctomi/.cd    labbits,  3(!) 

pulmonary,  281 
acute,  351 

relation  of  ions  to,  28 


Edema,  renal,  345,  348-350 

special  causes,  348-352 
Edematous  fibroma,  428 
Eel  serum,  177,  229.  318 

poisons   in,    164 
Eels,  vinegar,  376 
Ellusion,  tuberculous,  433 
Egg  yolk,  287 

Egg-albumen,   35,   192,   258,   353 
Eggs,  585 

Egyptian  mummies,  411 
Elirlich's   conception   of   nature  of   tox- 
ins, 128 

theory     of     toxins     and     antitoxins, 
177-178 
Eikosyl,  514 
E'laperine  snakes,   148 
Elapina-,  14!) 

Elastic  tissue,  degeneration  of,  371 
Elasticity,  decreased,  342 

of  young  tissues.  371 
Elastln,   419,  424 
Elastometer,   348 
Electric  charges,  28,  223,  379 
of  enzymes,  55 

current,  ell'ect  on  leucocytes,  259 
ell'ect  oh  motion,  378 
on  nuclei,  378 

shock,  cause  of  death,  378 
Electrical    conductivity,    27,    371,    514, 
527 

injuries,  253 

phenomena  of  colloids,  38 

resistance,  370 
Electricity,       efl'ect       on       protoplasm, 

378 
Electrolytes,  27-29,  32,  35,  336,  355 
Electro-negative  proteins,  396 
Electropism,  256 
Eledone  moschata.   170 
Elephantiasis,    340 
Elimination.  523 

rapid,  245 
Ellagic  acid,  463 
Embolism.  325-327 

air,  326  327 

fat,  325,  413 
Embolus,  223,  293,  325-327 

of  pancreas  cells,  387 

lilacental,  535 
ICmliryonic  origin  of  tumors.  496 
Mmphyscmalous  gangrene,  389 
l'hnp\(Mna,  311,   355,   433 
Emu'lsin,  61.  63.  67 
Emulsion   colloid.  452 
ICmulsions,  35 
Endemic  goiter,  601 
iMideslinate,  ])rotoniain,  166 
Kndocarditis,  .322 

uremic,  531 
I''iido(  nidium,  439 
Kndolvsin,  265,  266 

bacteriolytic,   280 
Endoplasm  of  bacteria,   106 


INDEX 


685 


Endosarc,  2G0 
Endothelial  cells,  219 
death  of.  .-^43 
hirjii",  273 
l)luiix<><"ytic.  2!)(! 
sudaiiopiiilc  clianjip.  272 
Eiulotlu'liolytic    scniiii,    23!) 
Eiulotlii'lioiiiii,   432 
Endotheliotoxii'  action,  293 
Eiulotlu'liotoxin,  153,  240 
Endotoxins,  103,  120.  129    130 
antitoxin  for,  130 
hacterial.  118 

in  ]>atlioL;(Miic  bacteria.   119 
Enterojrcnous  cyanosis,  580 
Kntorokinase,  61,  385 
Enteroliths,  463 

Enzvnies.    34.    40.    53-105.    280.    358. 
500   502 
amyloid,  422 
autolytic,  67 

of  tissue  cells.  277 
hartcrial.   113-120.  270.  3S2 

ininmnity  against,   117 
'    proteolytic,    resemblance    to    tryp- 
sin, 116 
relation  to  pathooenicitv,  116 
rrde  o*,   in  infectious  diseases,   110 
blood,  201 
dcaminiziiiir.    50] 
effect  of  light  on.  374 
electrical   charges,   55 
ethyl  bntyrate.  77 
glycolytic,   76 
hydrolvzing  fibrin,  282 
in  brain.   78 
in   kidney.   250 
in  venoms,  150 
inorganic,  56 

intracellular.   68-105.  264.  296,  368 
lecithin,   in   tissues,  77 
letu'ocvtic,  276 
lipolytic.  77 

in  lymphoid   cells.  77 
oxidizing.  68.  60.  371 

effect  of  liyhi  on.  375 
peptid-splitting,    361 
jicptolytic.   01 
protein   nature.   54 
protenlytic.  280.   315 
enzymes  of.  94-96 
of   leucocytes.    277 
purine.   497-498 
reducing.    74 

relation  of  antitoxins  to.  180 
of  complement   to,  211 
to  toxins.  59,  126 
resemblance  of  toxins  to.  67.   68 
specificity-  of,  59 
sugar-splitting.    55 
synthesis  by.  57 
toxicity  of.  61-63 
uricolvtic,  501.  633 
Eosin,   375,   493 


K(jsiMophil('s,  76 
Eosinoi)hilia,  200,  363,  417 

in  relation  to  glycogen,  432 

parasitic,    relation    to    anaphylactic, 
135 
5Jpeira  diodcnia,   159 
E|)idemic  meningitis,  355 
Epidermis,     superficial,     decomposition 

of.  412 
Ei)ididyniis.   474 
Epilepsy,  3(i0.  562,  586 
Ejjileptic   conxulsions,   562 
Epinephrin.   472,  576.   609 

arterial   dcgcncratidii    t'roni.   612 
Epi]>hanin    reaction.   209,   50S 
Kpisplenitis  scars,  424 
Kpithclial   hyalin,  424-425 

nuicin,  427 

pearls.  438 

])roIifcration  from  dyes.  493 
Epithelioid   cells,  273 
E])itheliolvsin.  241 
E]iitheli(.nia,   432 

cntaneous,    516 
Epitheliinn,   ciliated,   238 

renal,   calcil'cation   of,   438 
Equilibrium,   33,   56 

nitrogen,  506 

osmotic,  335 
Erepsin.  81.  358,  568 
Ereptase,    104 
Ergot,  576 
Erotism,    615 
Erucacic  acid,  224 
Erysipelas,  276,  321 
Erythema,  349 
Erythrocytes.   70,   289,  301 

infected,  474 

resistance  of,  230 
Erythrocytohsis.    214,    215-234.     See 

also  Hemoli/sis 
Erythrolvsis.  218 
Esterase,'  77,   408 
Ether,  50 

liyer  necrosis,   325 
Ethereal  sul]>hates,  506,  575 

sulphuric  acid,  5.  _ 
Etiiyl   alcohol.  57 

butyrate.   57 
enzymes,  77 

mercaptan.  567,  582 

sulphid,   567.  582 
Ethylidendiamine.    567 
Eu^lobulin.  181.  236.  290.  351.  353.  356 
Exanthemata,   aoite.  322 
Exophthalmic   -joiter.   66.    79.   231.   241, 
445,  604   608.  612 
iddiii    content.   605 
parathyroids  in.  607-608 
jdiospliorus  in.  605 
serum   treatment.   605 
Exophthalmos,  606 
Exosmosis.  350 
Explosion  of  fulminate.  380 


6S6 


INDEX 


Extirpation  of  organs,  71 
Extracellular  lysis,  207 
Extract  of  crayfish,  332 

of  leeches,  318.  332 

of   mussel,    332 

of  oysters,  332 
Extravasated  blood,   295-297 
Extravasations,    47S 

Extravascular  ])ressure.  decreased.  342 
E.xudates,  04,  93-94,  331,  433 

carcinomatous,  364 

cholesterol  in,  25 

inllanimatory,  331,  569 

l)neumonic,  404,   415 

putrid  jiurulent,  574 

tuliorculous.  358,  364 

turpentine.  358 
Exudation  of  plasma,  253 
Eyelids,  423 


Factors     ■which     influence     supply     of 

sugar  to  kidnevs,  637,  638 
Fat,  23-25,  33,  84,  127,  278,  279,  335, 
383 
accumulation,  pathological,  399-400 
bacterial.    110-112 

and   fatty  acids.    111 

effect  of  media  on.   111 
changes,     anatomical     and    chemical, 

402-404 
content,   in  organs,  403 

of  bile,  403    * 
decomposition  of.  products  of,  584- 

585 
embolism,  325.  413 
foreign,  de])o;^ition  of,  399 
formation,  bacterial,   399 

by  fungi,  399 
from  carbohydrates  in  food,  308 
from  chyle.  412 
from  fat  in  food.  398 
from  proteins  possible,  398 
in  bile,  326 
in  blood.  326 
in  caseous  material,  383 
in  chemical  comliination.  401 
in  fattv  organs,  analysis  of.  401 
in  urine,  326 
increase    of,    in    dc^cMicrating    organ. 

402 
infiltration.  397 
invisible,  made  visible.  403 
iodin    comiiounds.   400 
masked.   401,   409 
metabolism  of,  58,  602 
necrosis,  62.  80.  384-388.  414.  442. 

4S6 
nculrul,   migralinn   of.  412 
of   tulK'rcle  bacillus.    110    111 
solubility  of  dves.  50 
staining   in    tubercle   liacilhi.--.    1  1  I 
stains.  401 
sugar  from.  663 


Fat  tissue,  474 
Fat  droplets.    49 
Fatigue,   561-562 
mental,   562 
toxins  of.  75,  561-562 
Fatty    acids.    100,   278,    407,   441,    567, 
580 
crystallization  of,  412 
(Ives,   414 
free,  280.  401 
in  tubercle  bacilli,  112 
pathological  occurrence,  414 
soaps  of,  384 
toxicity  of,  414 
volatile,  280 
degeneration,  79,  397 
due  to  poisons,  407 
infiltration,    407 

metamoi'phosis,    75,    301,    328,    397— 
399 
by  absorj)tion,  408 
causes   of,   406-410 
relation  of  lipoids  to,  404-406 
reaction,  508 
substance,   303 
Fecal  elimination,  defective,  506 
retention.  524 
stones.   403 
Fermentation.    570-589 

of  carbohydrates,  products  of.   583- 
584 
Fermentoid,   120 
Ferments,  defensive,  205 
digestive,  506 
fibrin,    278 
glycolytic,  60 
protective,   197 
Ferratin.  479 
Fertilization,  285 
Fetal  tissue,  428 
Fetus,   intoxication    from,    536 

retention  of.  560 
Fever.  83.  322 

acetonuria  in,  500 
acute,    390 
Fibrin,  290 
calculus,  459 

content  of  blood  in  disease.  321 
enz\ines   hvdroiv/.ing,   282 
ferment.  67,   10.5.  278,  316 

thrombosis.   325 
formation,  315-317 
calcium    in,    310 
theories  of.  316 
ludrohsis.    509 
Fil)rinogen.  181.  290.  295.  315,  351,  353, 
356 
formation  of.  315 
Fibrinolvsin.   117 
Fil)rinolysis.  92.  315.  320.  325 
{•"ibrinous  thrombi.  .323 
{'"iliroid  gloMHMules.  424 
uterine.  510 

degenerating,  498 


INDEX 


687 


Fibroma,  432 

edeiiiutous,  428 
Filaria,   143,  340 

niediiieiisis,   14;? 
Fillration,  337 

theory    of    l\iiiiili     I'm  maliDii,    331- 
332 
Finson   li<^lit.  4S1 
Fisli,    lK)i^^uIl(m.s.    162    164 

plomaiiis,    1()3 
Fistula  biliary,  358 

Eck's,  520 

pancreatic,  443,   558 
Fixation  in  organs,  245 
Flagella,  112 
Flies,   black,    KiO 
Fluid,  distribution  of,  345 

migration  of,  412 
Fluids,  normal,  diil'erences  in,  353 
Fluorescent  substances,  375 
Fluorides  fixed  to  bones,  246 
Foam    structure,    268 

hypothesis  of  protoplasm,   51 
Foamy  phagocytic  cells,  405 
Food,  fat  from  fat  in,  398 

intoxication,  124 

poisoning,    122 

supplies,  338 
Foreign  body  giant-cell,  272 
organic,  438 

proteins,    165 

serum,   322 
Formaldehyde.   293 
Formalin,    43 
Formic   acid,   567,   583 
Fragility,  congenital  of  bones,  447 
Freckles,  467 
Free   acids,    247 

alkalies,    247 

fatty  acids,  280.  401 
Freezing,  elVect  of,  373 
Freezing-point,    depression    of.    of    col- 
loids, 38 

of  blood.  301,  308 
lowering  of,  50(i 
of  cerebrosiiinal  fluid,  360 
Friedlander's  pneumobacillus,  258 
Frog  poi.sons,   150 
in  skin  of,   162 

red  corpuscles  of.  229 
Fructose,    655,    656 
Fuchsin,  493 

bodies.    424 
Fungus,  fat  formation  by.  399 
Fusible  calculi,  458 


Galactose,  654 
Gall-stones,  448.  449 

calcium  oxalate.  450 

common,  449 

formation  of,  450-454 

relation  to  infection.  451 
Ganglion-cells,  474 


Ganglion-cells,  calcification  of,  438 

cortical,  chromatolysis  of,  531 
Ga.ngrene,     122,    373*,    388  390,    574 

carbolic  acid,  378 

dry,    388 

emphyseniat(jus,  3S'.l 

moist,  388 

of  lungs,  281,  389,  414 

ic-ray,  376 
CJjirtner  l)acilli.    17.'! 
(las  exchange.  302 

poisoning.   5tJl 
Gas-])roducing      organisms,      anaerobic, 

3(>3 
Gastric   carcinoma,   03 

fermentation,  457 

juice,  acidity  of,  245 
Gastro-intcstinal  asthenia.  614 

autointoxication,   566-589 

diseases,  75 

infections,   560 
Gaucher 's  disease,  405.  417 
Gelatin,  20,  36,  41,   194,  258,  319 
Gelatinase,  115,  280 
Gels,  34 
Generative  system,   relation   of  adrenal 

coVtex  to,  608 
Geometrical    structure,    relation    of,    56 
Geotropism,  256 

Germicides,   effect    of    alcohol    on,    28 
Giant-cells,  273,  416.  631 

foreign  body,  272 

formation  of,  264,  273-274 

of  tubercles,  76 
(Jlla  monster.  155 
Glabrificin,  185 
(Hand  decomposition,  412 
Glandular  secretions,  331 
(iliadin.  194,  286 
(Jlioma.  432 

(Jlobin.  20,  290.  295.  476 
(ilobulins.  22.  35,  64,  195.  289 

cell,  coagulation  of.  373 
<nomerules.  fibroid,  424 
(;  Incase.    291 

(Uucosamin,  110.  281.  426 
Glucose.  249.  639.  640,  642,  643 

injections.   642 
Glucoside,  226.  243 

as  antigen,    167 
(JIucothionic   acid.   279 
Glutamic  acid.  584 
( JIutaiiiinic  acid.  495 
(ilut  in-casein.  258 
Glycerol.  257,  407,  567 
Glycine.  286 
Glyco-alkaloid.  227 
GlycocoU,    195,    250.    258.    626,   630 
Glycogen,    25,    34,    84,    134,    246,    278, 
279,     357,     471,    496-497, 
544 

eosino])hilia  in  relation  to,  432 
graiuilcs,   49 

in  leucocytes.  432-433 


688 


INDEX 


Glycogen      in     patlioloiiical     processes, 
428-434 

in  tumors.  431-432 

inliltration   in  diabetes,  434 

pathologic    ofcurronce,   431 

physiological  occurrence,  429 

properties   of,   42*J 

relation   to  malignancy,  4!)G 
Glycogenolysis.  (514 
Glycogenolytic  ferment,  105 

secretion,  015 
Glycolytic  enzymes,  7G 

ferment.  GO 
Glj'conucleoiJroteins.   109 
Glyco-proteins,  21,  2;? 
Glycosuria,       G(l7.       (ilG.       (SM.      639, 
640,   657   659 

alimentary,  057 

relation  of  ions  to.  28 

specilic    meclianisms    by    wliieli    pro- 
duced. G40 
Glycothionic   acid,   422 
Glycuria,  636 
Glycuronic   acid,   245.   249 
Glycj'Iglycine,   20 

Glycyl-tryptophane  test  for  cancer,  501 
Goiter,  adenomatous,  GOl 

chemistry  of,  599-601 

colloid,  iodin  content.  600-601 

CA'stic,    tJOl 

endemic.  GOl 

epidemic  infections,  601 

exophthalmic,   Ctd,   79.    231.   241,   445, 
604   608.   012 
iodin   content,   005 
parathyroids   in,    607-608 
phosphorus   in,   005 
serum  treatment,  005 

hyperjilastic.   iodin  content,  601 

parencUx  matoiis.   001 
Gonorrheal   artlirilis,  433 
Gout,  31(1,  628  634 

calcium,  439 

chronic,  628 

guanine,  in  swine,  630 
Gouty  deposits,  405 

metabolism,  029 

tophi,  031 
Gram-positive   cocci,   20/ 
Gram's  staining,  inicleoproteins,   113 
of  bacteria.  112-113 
relation  f)f  cell  wall  to.  113 
unsaturated  fattv  acids.  113 
Granulation   tissue,   2G1,  425 
Grapes.  457 

<!r(i\ving  tissues   in    vitro,  274 
Growth.  253   288 

and  re|)aii-,  clicmical  basis.  285-288 

efFect  of  thyroid  on,  591 
fiuaiacol,   217 
Guanasp.  85.  497.  023 
("Juanidine.   142 
Gmiiiim\  019,  022.  023 

gout  in  swine.  03(1 


Guanosine,   622,   623 
Guanosine-deaminase,  623 
Guanylic  acid,  622 
Gums,  34 

Haptupjiokk,  67,  128,  177,  212 

group,    120 

precipitin,    191 
Ilay-fever,   147-148 

an   anaphylactic   reaction,    148 
Headache.  oS') 
Heart,  423 

disease,  valvular,  355 

hypertrophy  of,  284,  611 

murmur,  anemic,  431 
Heat  adaptation,  372 
Hedera  helix,  227 
Heliotropism,  255-256 
Helvella   esculentia,    146,  227 
Helvellic    acid,    227 

Hemagglutination,      by      lipase      from 
Ij-mphocytosis,  223 

by  vegetable  poisons,  223 

non-specific,  183 

separation  from  liemolvsis,  223 
Hemagglutinin,     153,     222-224,     240, 

325 
Hemangioma,  432 
Hematin,  290,  295,  477-478 
Hematogenesis,   231,   304 
Hematogenous  jaundice,   302 
Hemato-hepatogenous  jaundice,  230 
Hematnidin,    389,    296,    472,   478 
Hematoma.   415 
Hematoporphyria.   481 
Hemato]iorpIiyrin,   375,  480-481 
Hematuria  iu>onatorum.  033 
Hemicellulose,  109 
Hemochromatosis,   482-484 

local,  483 
Hemochromogen,    290,    295.    476 
Hemofuscin,  482 

Hemoglobin,  21,  35.  217.  229,  289,  290. 
302.  476-477 

anaphylactic    dill'erentiation    of,    175 

excretion,  230 

infarcts,   227 

metal>i)lism   of,   477 
Hcmoglobineinia,   230,  305 
Hemoglobinuria.     225,    227,    228,    230, 
305,  503,  477 

malarial.   130 

paroxvsnial.  232-233,  302 
Henndvinph  glands.  232,  290 
Hemolysin.    128,    153,   240,   358 

(piantitative  action.  221 
Hemolvsis,  31,  (i2,  207.  215-234,  487. 
5(i3 

action  of,  on   stiiuiia.  217 

acute  toxic,  478 

by  chemical  agents,  216 

by  phagocytes.  230 

bv  ]di\sical   means.  210 
bv  serum.   218   219 


INDEX 


689 


Hemolysis  by  tissue  extracts,  218 
by  vegetable  poisons,  225-226 
by  veuonis,  228   229 
in  burns,  2.52 
in  cancer,  504  .")(!,") 
in  disease,  229   232 
in   poisoning,   -'.i2 
inliiliition  ol,  217 
ineclianisni   of,  21.3 
pathology  of,  233-234 
postniorieni,    224 
serum,  made  of,  222 
splenic,   232 

variable  resistance  in   disease,   217 
Hemolytic       amboceptors,       219-221, 
232 
])roperties  of,   211) 
anemia,   230 
antibody,  218 

in  in  vitro  tissue  cultures,  170 
cancer    extracts,    492 
complement,  21'J,  221-222 
jaundice,    congenital,   231,  489 
lipoid,   307 
poisons,    177 

substances  in  tiunors,  504-505 
Hemophilia,  297-300,  322 
coagulation  time  in,  2'J9 
icterus,   487 
local,  298 
Hemopyrrole,  468 
Hemorrhage,   293-297,   305 
blood  changes  after,  294 
by  diapedesis,  294 
death  from,  390 
from  asphyxial   changes,   294 
metabolic  changes  alter,  301-302 
Hemorrhagic  ascites,   490 
cysts,  42G 

infarcts,  328,  611,  633 
nephritis,  385 
Hemorrhagin,    154,    240,    293 
Hemosiderin,  296,  329,  478-480 
Hemotoxins,  153 
Hepatic  necrosis,  371 
Hepatolysins,  238,  241 
Herbivora,  459 
Hernia,  433 

strangulated,    341 
Heroin  tolerance,  244 
Herpes  zoster,   276 
Heterocyclic  compounds,  470 
Heterolysis,  90,  328,  500 
Heteronephrolvsin,  240 
Hexoses,  22,  25,  650-654 
H  g  Cl„,  451 
H-ion  concentration,  236 
Hippuric  acid,  250,  525 
Hirudin.  319 
Histamine,   198.  576 
Histidine.  96,   100,  285,  544.  576 
Histon,   282,   309 
nucleinate,  494 
sperm,    177 
44 


Histon  thymus,  419 

llisloretention,  350 

llodgkins  disease,  71,  312 

llog-clioiera  bacillus,  324 

Homogentisic  acid,  73,  473,  577,  578 

Hordein,   194 

Horniouo,    disassimilatory,    593 

Hornets,   100 

Horniiication,  432 

Hyalin  casts,  425 

connective-tissue,  423-424 
degeneration,   3b2,  423-425 
epithelial,  424-425 
relation  to  amyloid,  424 
sunstance   of   Itovida,   279 
thromlii,  224,  324-325,  374 
Hydatid  cysts,  432 

fluid,   chemistry  of,   138 
wall,    138 
Hydrated  oxide  of  iron,  34 
Hydration,  colloidal,  347 
Hydremia,  293 
Hydremic      pletliora,      298,    334,      342, 

347 
Hydroa  aestiva,  481 
Hydrocele   fluid,    359,   415 
Hydroce])halus,  360 
Hydrochinon,  575,  577 
Hydrochloric  acid,  336 
Hydrogels,  34 
Hydrogen,  567  ' 

arseniuretted,    485 
sulphide,  245,  567,  581 
Hydrolysis,   19,  33,  429,  478,  569 

fibrin,  569 
Hydrophilic  colloids,  336,  409 

tendencies  changed  by  enzymes,  336 
of  colloids,  343 
Hydrophina?,  148,  149 
Hydrothorax,  356 
Hydroxy  1,  21 
Hydroxypurines,  287 
Hydro.xypyridene,  287 
Hydroxy-stearic  acids,  410 
Hypercholesterolemia,   417,    453 
Hyperemia,  312-315 
'active,  312-313,  341 
local,  253 
passive,  313 

blood   changes   in,    313 
pulmonary,  341 
Hyperemic  pressure  on  lymph  channels, 

340 
Hyperirritability,  531 
Hypernephroma^     432,     492,     497,     498, 
516,  517-518 
resemblance  to  adrenal,  518 
Hyperopnoea,  nocturnal,  500 
Hyperplastic  goiter,  iodin  content,  601 
Hyperpyrexia,  563 

Hypersensitiveness  of  nerve  cells,  530 
Hypersensitization,  169 
Hyperthyreosis,   71 
Hyperthyroidism,  604 


690 


lyoEX 


llyportrophy,  394 

adrenal,    532 

of  heart,  284,  611 
Ilypcealcification.  598 
H\i)()i)ln'seet(>mv,  ()]5 
Ilypopliysis.  42'(i,  4!)3,  614-616 

anterior  lolie,  014-015 

defects   in,   015 

posterior    lobe,   615 

]nuu'tiire  of,  615 
Hypopituitarism,    015 
Hypotliyreosis,  71 
Hyijoxanthine,    4HS.    (i]!i.    023 
Hysterical   vomit iiiii',  .')(iO 


IcHTiiyoToxiN,  104,  229 
Icterus.     See  Jaundice 
Idiojiathic  peritonitis,  353 
Idiosyncrasy,  243 
llhiminatin<i-  i^as,  70 
imliiliition,    3ti 
Iminazolyl    <;r()up.    015 
Immune  ainl)(>c('ptor.  211 
bodies,  210.  358 

reaction        inlluenced        by       electric 
charges,    175 
specificity  of,    171-177 
Immunity,  277 

against  bacterial  enzj'mes,  117 
malaria,    130-137 
non  antigenic  poisons,  243 
phytotoxins,  145-146 
]>rotozoa,    135 
toxins,   128 
cellular,  244 
natural,   178 
reactions,  chemistry  of,  165-209 

lipoids  in,    16S 
relation  of  ions  to,  28 
Immunization,   07,   178 
against  arsenic,  247 
therapeutic,   173 
Imjiaired  absorption,  246 
local  nourishment,  341 
Inanition,   560 

Indicanemia,  .■»02,  529,  573,  574 
Indi'^o  calculi,  45S 

Indole,   282.    40!l,   471,   507,   572,   573- 
575 
acetic  acid,  572 
propionic  acid,  571 
toxicity    of,    574   575 
Indole-acetic  acid,   575 
liuloh'iie,  248 

Indoplienol  reaction,  7(i,  248 
hidoplienoloxidase,    72 
hwlowi,  249,   572 
Infant  ih'  marasmus,  558   559 

s.Mirvv,   287 
Infantilism,  590,  603 

intestinal,   587 
Infants,  coairulation     time  oi  blood  in, 
322 


Infarction,  327-329 

hemorrli.igic.   Oil 
Infarcts,  76,  91,  97,  276,  281,  327,  308, 
431,  438,  441 

anemic,  327,  381,  407,  415 

choii'stcrol  in,  25 

hemoglobin,  227 

Jiemorrhagic,   328,   033 

iinman,  328 

old,  415 

splenic,  328     • 

uric-acid,  456,  627,  633-634 
Infected  areas,  481 
infection,    455 

Infectious  diseases,  320,  610 
acute,    317,    595 

role  of  bacterial  enzymes  in,  119 
Infiltration,  fat,  397 
Iniiamed  areas,  431 
Inllammation,  05,  253-288,  452 

acute,  361 

catarrhal,   427 

clironic,  273,  405 

of  l>ile  tracts,  453 

pulmonary,   405 

relation  to  chemical  alterations,  253 
Inllammatory     edema,     253,     340,     343, 
350-351 

exudate,  331,  509 
Infusoria,  130 

Inhibition  of  bacteria,   102 
Injection  of  glucose  solutions,  642 
Inorganic  en/\ines.   50 

poisons,  246-248 

salts,  28,  48,  51 
Inosine,  623 
Insects,   sting  of,  351 
Insoluble  proteins,  23 
Intercellular  edema,  337 

snl)stance.  51 
solution  of,  343 
Intermediary  metabolism.  524 
Internal  organs,  247 

secretion  of  tumors.  502-504 
Interstitial   nephritis,   349.   Oil.  (i27 

chronic,   527 
Intestinal  concretions,  462-463 

infantilism,    587 

mehinosis,  470 

obstruction.    433.    532,    580 
acut.-,  588   589 

])utrefact  ion,    245,    588 

sand,   403 

\vornis,  03.  432 
Intoxication,  050 

acid,  292.  534,  547-550,  584 
non-diabetic,  557-561 

anaphylactic,   (i2 

from   fetus,  5.30 

uric-acid,  (127 
Intracai)illa)y  blood  ])rcssui-c,  334 
Intracellular   edema,    ■3.")7 

enzymes,    68    105,    204,    290,    308 

lysis,  208 


INDEX 


691 


Jiitraci'lluliir  oxidasos,  4U7 

parasite,  47-4 
liilraliepatic    necrosis,    387 
liiliali^iameiitary  papillary  cysts,  514 
Jntravitain   cell  autolysis,  408 
liuilin.   JSt) 
liivertase.    lil,    l&I 
Invertebrates,    IDj 

shells  of.  4;J7 
Invertin,    07 
Jnvdliitioii  of  uterus,  !)2 
Iodides.  71.  2!)3 
lodin,  (iOU 

compounds    of    fat.    4()(» 

content     of      colloid     goiter,      600- 
601 
of  exoiiiitiialniic  goiter,  OO.j 
ot  hyperplastic  goiter.  GUI 
Iodoform,  •24;5,  270 
lodophilia,  4.5.S 
Ionization.  27-29 
Ion-protein.  26 

compounds,   28 
Ions.  20 

antagonism  of,  247 

relation  of,  to  diuresis,  28 
to  edema,  28 
to  slycosuria,  28 
to  immunity.  28  ■ 
Iron,   20,  45,   258,   303 

hydrated  oxide  of,  34 

in  tumors,  500 

starvation  cause  of  chlorosis.  304 
Isoagglutination,    224,    231 
Isoamylamine.  507 
Isocetinic   acid,    110 
Isohemolysins,  220 
Isoneplirotoxins.  240 
Isotonicity,  303,   305 

Japanese  lacquer,   409 
Jaundice,   71,   218.   220.    227.    203.    .305, 
321.  410.  478.  484   491.  507 

congenital  hemolytic.  231,  489 

dissociated,  48!) 

etiology  of,  484  486 

gravis,  488 

hematogenous,  302 

heniato-hepatogenous,  230 

hemophilia.  487 

in  newborn,  480 

local,  484 

necrosis.   487 

obstructive.  231.   410.   480 

pigmentation  in.  489-490 

relation  to  infections.  488  , 

Jecorin.  100.  270,  200 
Jequirity.   257 

Kauyokixests.  45,  285 
Karyolysis.  368  369 

luiclear  chanires  in.  360 
Karvorrhexis.  218.  250.  .328.  368-369. 
380 


Kalaljolic   [iroccsses,   371 
Katabolisni,   normal,   200 

nuelein,  310 

protein,  increased,  .302 
Kenotoxin.  501 

Keratin,  40,  285,  424,  408,  510 
Keratoiiyalin,   425,   402 
Keratitis,  250 
Ketone,  cyclic,  530 
Ketoreductase,   74 
Kidneys,    480 

amyU.id.   404,   415 

atrophy  of,  474 

concretions,  amyloid,  423 

cn/.ynie  in,  250 

large  white,  405 

lipins  in,  23 

sugar  sup])ly  to,  637 
Kinase,  01,  501 

Klausner's  serum  reaction,   237 
Krait  venom,  155 
Kupfl'er  cells.  474 
Kynurenic  acid,  572 

Laccase,  07,  73 

Laccol,  73 

Lactic  acid,  85,  248,  257,  278,  047.  648 

relation  to  drug  jjoisoning,  556 
Lactim,  020.   028 
Lactose,  280 
Lactosuria.  656 
Laking  of  blood,  215 
Lamprey  serum,  toxicity  of,   104 
Lanthanin,   44 

Lardaceous   degeneration.   417 
Large  white  kidnej^s,  405 
Larynx,  423 
Lassitude,   585 

j^athrodectes  tredecim-guttatas,    158 
Laurie  acid,  111 
Lead.  240 
Lecithids,  228 

Lecithin.  23,  24.  25,  85,  03,  100,  217, 
270,  285,  280,  303.  357,  401, 
437,  401.  502,  000 

as  antigens,  236 

decomposition.  584 

enzyme  in  tissues,  77 

in    diphtheria    bacillus.    Ill 

relation  to  opsonins.   160 

stearvlolevl.  24 
Leech  extract.  318.  332 
Leptothrix.  405 
Leucine,  20.  87.  06.  258.  270.  309.  327. 

357.  380.  542 
1  eucocidins.  230 

Leucocytes.  70.  75.  88,  01.  103,  238.  280. 
290,  404 

action  of  thorium-x  on.  377 

autolysis  o^  308 

chemotaxis  of,  250 

dispersion  of.  273 

e'lect  of  electric  current  on.  250 

glycogen  in,  432  433 


692 


INDEX 


Lenoocytos,  migration  of,  253 

due   to   c'liaiige   of   surface   tension, 

271 
relation    of    cell    types    to,    259- 
261 
phagocytic.   631 

action,    253 
proteolytic  enzymes  of.  94-96,  277 
relation  to  ameba.  2G() 
thermotaxis  of.  261-262 
Lencocytic  enzymes,  270 
protease,  (i3,  6.1,   103 
walls,  272 
Leucocyto-agghitinin,    240 
Leiicocytolysins,    1.53 
Leiicocytolysis,  214 
Leucoc'vtolvtic  serum,  239 
Leucocytosis,  239.  292,  319,  375,  450 

general,  2 J '3 
Leucocytosis   in  burns.  261 

mastccU,   261 
Leucocytotoxins,   154,  239,  240,  311 
Leueolvsis,  218 
Leucoiienia,   200,   239,  320 
Leukemia,    71,    75,    70,    103,    293.    298, 
307-312,     321,     376,     450, 
031.  034 
chemistry  of  blood  in,  308 
eliroiiic.  521 
lymi)hatic.  260,  307 
myeloid,  433 
myelogenous,  95,  307 
protein  metabolism  in.   309 
a-ray,  377 
Leukemic  blood,   coagulation  of,   308 
Leukoprotease,   95,   277,   278 
Lerulose,    655-656 
I^evulosuria.   031") 
alimentary,  055 
idiopathic.  056 
mixed.  656 
spontaneous,   656 
alimentary.   655 
Light,  bactericidal  action  of,  374 
dillVrent   rays  of.   374  ^ 
effect  on   antibodies,  374 
on  enzymes.  374 
on  oxidation  processes,  375 
on  oxidizing  enzymes,  375 
on   ]iroteiti  solubility,  375 
on    tissues,   374   377 
on  toxins,  375 
stroke,   481 
Lime  laden  drinking-water,   453 
Linin.    44,    45 

Lipase,    58,    62,    67,    77-80,    95,     103, 
138,  262,  278.  280,  291,  358. 
407,  535,  595 
in    1\  inphocytes,   79 
in  ui'ine.  78 
pancreatic.    3*^6 
Linemia.  412   414 

in    iilc, holism.    413 
]>ipins.  23   25,  400,  498-499 


Lipochrome,   393,   474-476 

tests  ft)r,  475 
Lipoeyanin,   475 
_L,ipofuscins,   393,   475 
Liooidal   degeneration,   410 
Lipoidemia.   413 

Lipoids.  23-25,  65,  127,  243,  461 
adrenal,  in  arteiial  disease,  009 
in   pneumonia,  tiOO 
in  renal   disease,   6!)9 
anisotropic,  25 
doubly   refractive.   25 
echinococcus,   antigen    from.    108 
hemolytic,    307 
in   immunity  reactions.   108 
relation   t-,)  antiliodies,    169 

to  fatty  metamorphosis,  404-406 
tape  worm,   antigen  from,   168 
Lipolysis,    413 
Lipolytic  enzymes,  77 

in   lymphoid   cells,   77 
Lipoma."  432,  511-512 

calci'ying,  441 
Lipo-peptids.   401 
Liquefaction,  276 
necrosis,  382 
aseptic,  276 
Lithemia.  ()27 
Lithofellic  acid,  463 
Liver,  83.  486 
abscesses,  135 

acute  yellow  atrophy,  379,  403,   538, 
539-550.  557.  569 
l)lood  in.  546 
alcohol  oxidase  in,  248 
atroi)hy  of.  474 
autolysis  of,  231 

effect  of  thyroid  on,  591 
caffein  demethylated  in,  247 
chhu'oform   necrosis   of.   320 
cirrhosis  of,  320.  355 
degenerations,   100—102 
diseases,  292.  480 
edema  of,  340 
necrosis,  324 
ether,  325 
passive  congestion.  431 
site  of  urea  formation,  526 
Luml)ricin,  143 
Liuigs,  e<lema  in.  3  11 
gangreiu'  of,   414 
stones.   459.  464 
Lupus.  374 
Lymiih.  absorption  of.  338-339 

channels,  hvjiereinic   ]iressure  on.  340 
comiiosition  of.  330  331 
due  to  endotiielial  secretion,  332 
due  In  filtration.  3.31 
lldw.   iiostmortem.  .340 
formation   of.   331-338 
ell'ect    of   ai'ids   on.    337 
relati'-n  of  chvle  to.  331 
to  blood   i)lasnni,   330 
secretion.   ])ostnu)riem,  332 


INDEX 


693 


JvViMjili,    llnnaiic,   ojjiuolie    pressure   ol, 

331) 
LymiJliayogues,   332,  341) 

Lr\stuiloid,  :i'62 

t'liufl,  JOt) 

^llIlHllatlllg,  332 
J-viiipiiaiigK)iiia,   432 
J.vmpiiaiiyUi-s,   elirunic,   340 
Lymplialic  congeslion,  passive,  341 

R'liKeiiiia.  20U,   307 

oUslriiL-lioii,  a40-341,  348 

tissue,   4!I4 
Lympliatulytic  serum,  240 
Lymph-giauds,  TiS,   102 
Lympliue\  les,    7U,   272 

lipase  in,  'i  \) 

ui   lubereulosis,  200 
Lympliocytosis,  375 
Lyniplioiu  cells,  lipolytic  enzymes,  77 
Lymphosarcoma,  4!)4 
Lysiuogeii,   221 
Lysiiis,  Go,  100,   17it,  195,  285,  286,  544 

cancer,  241 
Lysis,  207 

extracellular,  207 

intracellular,  208 
Lysol,  245 


^lACRUPlIAGES,    265 

mononuclear,   I'JO 
Magnesium,  290,  303,  444 

salts,  eilect   of,  on  phagocytosis,   263 
Malaise,  ot)') 
Malaria,  136.  231,  321,  447 

immunity  against,   136-137 
^lalarial  hemoglobinuria,  136 

pigment,  46 i 

pigmentation,  474 
Malic  acid,  254 
Malignant  growths,  238 

pustule,   276 

tumors,   284,   569 

chemistry   of,   515-518 
Malmignatte,  158 
Maltase.   103 
-Maltosuria,  657 
Mammary  gland,  88 
-Manganese,  500 
Mania,  531 

Marasmus,  infantile,  558-559 
Margaric  acid,  crystals,  414 
Margarin  crystals,  389,  414 
Marsh-gas,  567 
Masked  fats,  401.  409 
Mast -cell  leukocytosis,  261 
Maturity,  delayed,  590 
Meat.  627 
.Meal-worms,  469 
Mechanical  atlinit}',   37 

injuries,  253 

stimulation,  263 
^Mechanistic   school,   330 
Medulla,  adrenal,  472 


Mcioslagmin  reaction,  208  209 

test,  c>08 
-Melanemia,  471 
Melanin,   ,.i,  207,  467-472,  577 

as  antigen,  170 

composiuon  ol,  468 

properties  of,  469 

loxicity  of,  471 

tumor,  496 
Melanogeii,  tej>ts  for,  470 
-Meuino-prolein,  4(»b 
AieUlno.•^arcoma,  469 
-Melanosis,  467 

intcBtmal,    470 
Melanotic  adrenal  tumor,  472 

tumors,  467,  471-472,  502,  577 
Melanuria,  4(0,  473 
-Melena  neonatorum,  298,  321 
Melituria,  636.     fcsee  also  Diabetes 
Membrane,  nuclear,  45,  47 

semipermeable,  30 
Meningeal  ellusions,  359 
-Ueningitis,  360,  361 

epidemic,  355 

tuberculous,   360 
Menstrual  blood,  320 
Mental  diseases,  229.  362 

latigue,  562 
Mercury,  246,  258,  276,  542 

bichloride  of,  43 

hxation,  246 
Metabolic  activitj',  335 

changes  after  hemorrhage,  301-302 

disturbances,    566-589 
Metabolism,  57,  58,  59,  S7,   171 

abnormalities  in,  523-565 

basal,  590.  602 

carbohydrate,  602,  635-671 
ell'ect  of  thyroid  on,  591 

fat,  602 

gouty,  629 

111  cancer,  505-507 

intermediary,  524 

of  hemoglobin,  477 

protein,  444 

in   leukemia,  309 

in  pernicious  anemia,  306 

j>urine.    629 

relation  of  thyroid  to,   590-591 

respiratory,  444 

uric-acid,  618-634 
Metalbumin,  513 
Metallic  coi)per,  258 

poisons,  246 

sulphides.    247 
Metal-protein  compounds,  247 
.Mctanu)r])liosed    calculi.    455 
-Metamorpliosis,     fatty,     75,     301,     328, 
397-399 
by  absorption.  408 
causes  of,  406^10 
relation  of  lipoids  to,  404-406 
Metaplasia.    285 
-Metastatic  calcification,  438^39 


694 


INDEX 


Metazou,  372 
:Motliiino.  2r)0 
Methemoploljin,  297 
^Metlicnioglobineniia,   481 
Methyl    cyanainide,    598 

guanidino,  505 

mercaptan,  5(17,  582 
Metliylation,  247,  250 
Mctlivk'iu'  blue,  reduction  of,  253 
Metliylfiuauidine,    199 
Micrococcus  cereus  tiavus,    132 
Migration  of  fluid.  412 

of  leucocytes,  253 

due   to   change   of   surface   tension, 

271 
relation  of  cell  types  to,  259-261 

of  neutral  fat,  412 
Mitochondria,  47,  49,  395 
Mocassin  venom,  150,  228 
]\Ioist  gangrene,   388 
Molecule,  colloid,  395 

radicals  in,  arrangement  of,  176 
Monobasic  urate,  G20 
Mononuclear    macrophages,    190 
^l()n(>]iotassium  phosphate,  561 
Monosddium   urate,  620,  629 
Monovalent  antigens,  179 
]\Iorbus   maculosus,   480 
Morchella  esculenta,  228 
Morner's   body,   309 
Morphine,   165,  244,  258,  379,  560,  592 

tolerance,  244 
Motile  bacteria,  256 
Mouse   tumors,    173,   432 
Mucin,   109,   195,  281,  356,  426 

connective-tissue,    428 

epithelial.    427 

in  myxedema,  603 
Mucoid  degeneration.  426,  427-428 
Mucoids  of  ovarian  cysts,  513 
Multiple  myeloma.  518-519 

sclerosis,  414 
Mummies.   60 

Egyptian,  411 
MurirnidiT?,    163 

Muscarine.   123,   124,   260,  567.  585 
^Muscles,   22.   83 

decomposition  of.  412 

waxy  degeneration,  392-393 
Muscular  asllienia,  614 

dystrophies.    393 
Musli rooms.  227 

poisons,  146,  177,  539 
Mussel,  extract  of,  332 
Myelins.  25,  404,  461 
Myelol)last8,  476 
Myelocytes,    476 

Myelof/enous    leukemia..    95.    307 
M'veloid    leukemia,    4.33 
ATveloma,   multiide.   518  519 
Myelopathic  al1)umosuria.  519   521 

occurrence  of.   520   521 
Myelotoxic  seiiim,  240 
Mykol,  111 


Myocardium,   486 

chronic  degeneration  of,  284 

lecithin  in,  24 
Mvogen-fibrin,  soluble,  391 
JMvoma.  432,  509 
:\Iyosin,  391 
Myosinogen,   391 
Myristic  acid,  514 
Myristinic  acid,  110 
Mvrosin,  61 
Myxedema.  428,  586,  590,  601-604 

mucin    in,   603 
Myxofibroma,  428 
:\rVxoma.  428 
Myxosarcoma,    428 

Naphthalin,  249 

Narcosis   chloroform.   540-541 

Narcotic    poisons,    87 

Narcotics,   246 

Nasal  septum,  423 

Nascent  oxygen,   60 

Nastin  as  antigen.   168 

Necrobiosis.  99,  117,  368,  438 

Necrogenic  substances,   135 

Necrosis,   253,   276,   367-390,   631 

anemic,  328,   368,   372,   381 

aseptic   liquefaction,  276 

causes  of,  371-381 

chloroform.  408 
of  liver,   320 

coagulation,    381-382 

due  to  physical  agents,  380-381 

due  to  ic-ravs,  376 

fat,  62,  80,"  384-388,  414,  442,  486 

hepatic,  371 

icteric,  487 

intrahepatic,  387 

liquefaction,   382 

liver,  324 
ether.  325 

radium,  376 
Necrotic    areas,    central.    273 

tissue,  97 
Necturus,  229 
Negative  charg(\  increased.  285 

chemotaxis,    257 
Nematodes,  140   143,  432 
Nephrectomy,  292 
Nephritic       transudates,       coiu])ositi(m. 

354 
Nephritides.  chronic,  240 
Nephritis,    70.    78.    231,    293.    208,    321. 
343.  416.  417,  558.  587.  610, 
631 

acute,  292.   349.   379 

autolysis  in.  98 

chroTiic.    349.   424.   527 
interstitial.  527 

liemorrhagic,   385 

interstitial.   349.   611.  627 

parenehvmatous,  349,  350,  527.  570 
clironic.    364 

uranium,  344 


iNimx 


695 


Xoi)liiolysins,  238 
Ncplirolysis,  a',i2 
Ni'Kluohtic   sc'ium,   240 
Xephrotoxins.   240 

Nerve    cells,    foajiulatiiij;    teiiiiiorature. 
372 
hypoisensitiveness  of,  530 

neuralpie,  351 
Nerves,  vasoconstrictor,  253 

vasodilator,    253 
Nervous  diseases,  58G 

system,  sympathetic,   472,   503 
Neuralgic  nerve,  351 
Neurine,    123,    124.   562.   507,   585 

toxicity  of.  124 
Neuritis,  287 
Neurotibroma.  472 
Neurofrlia,   4(il 
Neurolytic   scrum,    240-241 
Neuropathic   edema.   351 
Neurotic  edema,  345 
Neurotoxins,   153 

venom,    154 
Neutral   fat,  migration  of.  412 

phos])hates.    021') 
Neutralization.   ?4S 

by   chemical   comliination.   523 

of    organic   acids.    251 
Neutrophiles,   203 
Newborn,  jaundice  in,  486 
Nicoll  prisn;s,  404 
Nicotine,   012 
Nidus,    451 

Nile  blue  sulphate,  401 
Ninhvdrin,  204 

test,    301 
Nissl  bodies,  46,  49 
Nitrobenzol.  217 
Nitrogen,  326 

amino-acid,  520 

blood,  Jion-protein,  534 

colloidal,    500 

dianiiiio,  545 

e(|uilibrium,    506 

non -protein,    520 
Nitroglycerin,    217 
Nitro-proteins.   176 
Nocturnal   hvperopnoea.   560 
Non-electrolytes,  35,  330,  347 
Nonoic  acid.  414 

Non-protein  organic  contents.  357 
Non-specific    hemagglutination,    183 

reactions  in   typhoid   fever,   174 
Nuclear    membrane.    45,    47 
Niwlcase,  44,  85,  07,  105,  300,  501 

purine,    622 
Nucleic  acid.  22.  23.  616 
bacterial.  100 
thymus,   621 
Xiiclein.  22 

bodies.    610 

hydrolysis,  284 

katabolism,  310 
Nuclcinic  acid,  250 


Nucleo  albumins,    23 
Nucleohiston,   282 
Nucleohis,    IS,  45,  47,   48 
Nucleo-profeins,    21,    22,    44.    105.   221, 
238,  241.  270,  285,  280,  357, 
45(;,  632 

bacterial,   108 

impure,  318 
Nucleoside,    022 
Nucleotids,  622 
Nucleus.    18,   454 

acidity  of,  40 

chemistry   of,  44-46 

of   bacteria,    100 

structure   of,   48 

Obesity,  416,  500 
Obstructive  jaundice,  231.  416 
Occlusion  of  thoracic  duct,  340 
Ochronosis,   473,   482,   577 
Oil,  croton,  270 

of  turpentine,  276 

rancid,  in  alkaline  solution,  267 
Oleic  acid.   110,  22S 
Oleum    pulgeii,   400 
Ophiotoxin,   150 
Opsonins.   207-208.  264,  358 

action  of  chemicals  on.  208 

chemical  nature  nf,  208 

relation  of  lecithin  to,   100 
Orang-utan,   025 
Organ  extracts,  318 
Organic  acids,  346 

contents,   non-protein,   357 

dves,  34 

poisons.    248  252 
Organs  of  cancerous  patients,  105 
Ornithine,   581 
Osazone,    387 
Osmic   acid,  43 
Osmosis,  29  33.  337 
Osmotic    concentration.    284 

equilibrium.    335 

exchanges.  337 

pressure.  30.  32.  210.  332.  334-336, 
371.  305.  .527.  643 
changes   in.    380 
disparity  of.  344-346 
increase  of,  254 
of  colloids,  38 
Ossification,  defect  in,   617 

relation  of  calcification  to.  435-436 
Osteitis  deformans.  445 
Osteogenesis   imperfecta.    447 
Osteohemachromatosis.   474 
Osteoid  change,  285 
Osteoma.   432 
Osteomalacia.  438,  443  445.  446,  520, 

500 
Osteomyelitis,    117 
Osteoporosis,  500 

senile,  444 
Osteosarcoma.  438 
Ova.   285 


696 


IXDEX 


Ovarian   colloid,  513 

cystadenoina,  513 

cysts,  contents  of,  512 
dermoid,  514 
mucoids  of,  513 

tumors,  -42(5 
Ovaries,  a;-ray  atrophy,  37G 
Overproduction      vs     underconsumption 

in  diabetes  mellitus,  070 
Overton  theory,  31,  50 
Ovomucoid,  1!)5 
Oxalic   acid,   248,  251,   G20 

calculi,    584 
Oxaluria,  5S4 
Oxazone   base,    401 
Oxidases,  55,   71,  05,    103,  280,  358 

alcohol,   71 

in  tumors,  76 

intracellular,  407 

reaction,  309 

xanthine,  71 
Oxidation,  33,  68,  69,  248,  638,  639 

impairment,  345 

of  cystine  sulphur,  614 

processes,  374 

efl'ect  of  lights  on,  375 

reduction  in,  348 

uric   acid,   248 
Oxidative  processes,  69 
Oxidizing  enzymes,   68,   60,   371 

efl'ect  of  light  on,  375 
Oxy-acids,   101 
Oxvbutyric  acid,  567 
Oxygen,  326 

active,  374 

nascent,   60 

relation    to   sarcolactic    acid,   555 
Oxygenase,   72 
Oxyhemogloliin,    35,    477 
Oxyniandelic  acid,  543 
Oxypurines,    622 
Oysters,   extract  of,   332 

Palmitic  acid,  514 
Palsy.     See  Paralysis 
Pancreas,   80,  85 

activating  substance,  77 

atropliy  of,  393 

cells,  eml)oli   of,   387 

diabetes,  665   671 

historical,  (165 
symptoms,  667 

extiry)ation,   430 
effects  of,  6()5 
islet  theory,  666 

nature  of  inlernal  sccr(>tion  of.  666 

self -digest  ion   of,   388 
Pancreatic  aulolvsis,  386 

calculi.  461    462 

dial)etes,    412 

fistula,   443,   558 

juice,    127.   245,   542 

lipase,  386 
Pancreatitis,  385,  48(;,  6(t7 


Papain,  62,  103 

Papayotin,    258 

Papillary    cysts,    intraligamentary,    514 

Parabanic   acid,   626 

Parabiosis,  564 

Paracentesis,  356 

Parachroniatin,  44 

Paracresol,  571,  576 

Paraheinoglobin,  297,  477 

Para-hydroxy-phenyl-ethylamine,  576 

Paralactic  acid,  544 

Paralinin,  44 

Paralysis,  487,  531 

agitans,   599 

diver's,  326 

general,   360 

progressive,  301 

vasoconstrictor,  312 
Paramecia,  252 
Paramoecium,  266 
Paramucin,   427,   513 
Paramucosin,  513 
Paramyosinogen,  391 
Para-oxyphenyl  acetic  acid,   571,  576 
Para-oxyphenyl-propionic  acid,  571,  576 
Paraphenylene  diamine,  248 
Parasites,  animal,   260,  340,  432 
chemistry  of,  134-143 
eosinophilia  (chemotaxis) ,  134 

intracellular,  474 
Parasitic  eosinophilia,   relation   to  ana- 
phylactic, 135 
Parathyreopriva,    tetany,    598 
Parathyroid,  597-599 

in  exophthalmic  goiter,  607-608 

tetany,  549 

tissue,   241 

tumors,    432 
Paratliyroidectomy,  598 
Paratyphoid  bacilli,  173 
Parenchymatous   degeneration,    395 

goiter,    601 

nephritis,    349,    350,    527,    570 
chronic,   364 
Parotid,    (i03 
Parovarian  cysts,  514 
Parowsmal  hemoglnlunuiia,  232-233, 

302 
Pars  intermedia,   015 
Passive  congestion,  314,  333 
of  liver,   431 

hyperemia,    313 

blood    changes    in,    313 

lympliatic  coiigestiou,  .341 

sensiti/ation.   191,  203 
Pattiogenic  bacteria,  <'ii(loto\iiis  in.  119 
Pellagra,  287 
Pt'lvis,    renal,   457 

Pentoses,  22,   25,    ini.  497,   649.   650 
Peiitx)suria,    577 

chronic,  650 
Pepsin,  (il.  62.  04,  07,  103.  127.  5(i6 
P(>ptid-splitf  ing  enzymes.   .'iOl 
Peptolylie    enzymes.    91 


INDEX 


697 


r.'ptuncs,   27S,   27!l,   2Sl,   .iOi),  544,   'iiil, 

.HiS 
l*eii'fiita<ri',  l)li)(>d  sufjar,  (i41,  (i4'2 
Perilnoiieliial  {jlamls,  4ti4 
rericardial   fuKillcatitin,    4:50 
rericarditis,  355 

ureiiiic-,    ").!1 
Peritonitis,  71,  355 
carciiiuinatuiis,   353 
general,   433 
idiopathic,   353 
IVniiangaiiate   reduction   index,   361 
rerineability,    3i),    370 
tor  vital   stains.  371 
of  capillaries,  333-334 
lo  colloids,  371 
IVniicious  anemia,  231,  305-307,  317, 
477,  587 
analysis  of  orjjans  in,  305 
calorinietric   studies   in,   306 
causes   of,    307 
ciieniical  changes  in,   305 
due  to  hemolytic  poisons,  300 
iron   ill  corpuscles,  305 
l)rotein   nn'tabolism    in.    300 
vomiting  of   pregnancy,    538 
Peroxidase,   72,   595 
Peroxides,    GO 

Phagocytes,  hemolysis  by,  230 
Phagocytic   activity,   417 
cells,"  219 

foamy,    405 
cndotiu'lial    cells,    296 
leucocytes,    031 
Phagocytosis,    39,     91,    207,    208,    253. 
254  256,    261,    262-267 
273,  314 
by   protozoa,  202 
etfect  of   calcium   salts  on,   263 

magnesium  salts  on,  203 
inlluencc  of  serum  on,  264 
relation   to  cholesterol,  263 
results  of.  204 
theories  of,  266-275 
Phallin,    147,   227 
Phallusia    mamilhita.    170 
Phanerosis,  402 
Phaseolus   nuiltillorus,   223 
PluMiacetin.    4S2 
Plienol.   24S.   25S.   451,   507,   571,   575 

])ois()ning,   245 
Phenolase,  73 

PlK'iiylalanine.   469,   495,   507,   576,   578 
Plienyl-ethyl  amine,     576 
Philo'catalase.    70 
Phlclxdiths.  459 
Plilogosin.    250 
Phloretin.    059 
Phloretiiiie    acid.    059 
Phlorhizin,   500,  659.  (JCO 

diabetes,  493,  557,  659-664 
Phlorose,   659 

Phosphate     anunonio-magnesium,     455, 
456 


Phospliate  calculi,  457  458 

triple,  3H9 
Phosphates,  357 

neutral,  626 
Phospluitids       in       tubercle       bacillus, 

111 
Phosi)ho-glycn proteins,    23 
Phospholipin,    4!l 

protein,   310 
I'liospiio-nuclease,  022 
IMiospiioproteins,  23 
Phosphorescence,    380 
Phosphoric   acid,   20,    303,   441,   444 
excretion,   310 
in    calcitication,    442 
Phosphorus,    247,    252.    293.    399,    407, 
499 

in  exophthalmic  goiter,  005 

poisoning,  74,  100,  317,  320.  413,  539, 
540,  557,  569 
Photosensitizing  action,  480 
Phthisis,  79.  281, 
Physical   absorption,  24() 

chemistry  of  cell,  26-43 

solution,  246 
Physico-chemical  factors,   349 
Phvto-preciptins.    192 
Phytotoxins,  144-148.   107.  293 

immunity    against,    145-146 

relation  to  proteins,  144-145 

toxic  action,    140 
Pigment,  279.  295.  389 

bacterial,    132-133,   2S0 
alcohol  soluble,   133 
insoluble,   133 
water  solulile,  133 

bile.  486  487,  566 

blood,  476-484 

fornuition,  73 

malarial,  467 

urinary,  525 
Pigment-granules,  49 
Pigmentation,  73 

in  jaundice,  489-490 

malaiial,  474 

patliological,  467-491 
Pilocarpin,  260,  312 
Pineal  gland,  017 
Piper az in,  311 
Pituitrin,  014 
Placenta,  putrid,  574 

retention  of,  560 
Placental  cnd)oli.  535 
Plague  agglutinin,  1S4 

bacilli.   107 
Plant  tissues.   195 
Plasma,  exudation  of.  253 

fat  of,  290 

sugar  of.   290 
Plasma phaer(>sis,  295 
Plasmodia.  231 

Plasmodium  malari:c.  136-137 
PlasuKdysis.   107 
Plasmoptysis,  31,   107 


698 


INDEX 


Plasmorrhexis,  31 

Plasmose,   316 

Plasnioaomes,  430 

Plastein.  57,   105,   115 

Plastin,  44,  45,  49 

Platypus  venom,  157 

Pletiiora,     hydremic,     298,     334,     342, 

347 
Pleurisy.  281,  35G 

tubereulous,   350 
Pleuritic  fluid,  521 
Pluriunim  resistance,  305 
Pnein.  09 

Pneumoliacillus,  Friodlandor's,  258 
Pneuniocoecus.    70,    129,    198,   225,    289, 
482.  488 

anaijhvlatoxic  poison.   198 
Pneumonia.   03,  78,  281,  282.   292,  293, 
319,  321,  350,  355,  413,  431. 
433.  480.  500,  509,  034 

adrenal  lipoids  in,  009 

autolysis  in,  96 
Pneumonic  exudates,  404,  415 

sputum.  490 
Pneumonoknniosis.   465-466 
Pneumothorax,  chemistry  of,  365-366 

dry.  305 

purulent,  305 

putrid,  305 
Poison,  ants,   160 

bacterial.  592 

bee,  160 

toxolccitliidin    in.    100 

black  flies,   100 

blood,  400 

causing  somnolence.  502 

centipede.   159    160 

convulsive,  87 

crab,  164 

froof.  150 

hemolytic.  177 

hornets.   100 

in  coelenterates,  164 

in  eel  serum,  104 

in  skin  f)f  frf)fr.  102 
of  salaniandcis.    162 

inorr'anic.   246   248 

I'letallic.  240 

musliroom.    140,    177 

naicotic,  87 

non-antifrenic,  defense  apainst,  243- 
252 
immunity  a<;ainsl.  243 

orpanic,    248   252 

prodiu'cd    in    su])erficial   burns.   562 
565 

protoplasmic,  379 

scorpion.    157    158 

spider,    158    159 

steato-renetic.   403.    409 

toad,  150.  161-102 
in  blood,  101 

vcfretable,   hemolysis  by,   225   226 

wasps,  100 


Poisoning,  485 

abrin,  histologic  changes,  140 

aceto-acetic  acid,  553 

arsenic.  342,  539 

bichlorid,   532 

chloroform,  379,  539,  557 

chromium,  477 

cobra,    152 

drug,  relation  of  lactic  acid  to.  550 

food,    122 

gas,  501 

mushroom,  539 

])henol.  245 

phosidiorus.    74.    100.    317,    320.    413, 
539,   540,   557,   569 

ptonuiin,  sources  of,   122 

ricin,  245.  573 

histologic  changes,   146 

shelUisli,   342 

snake,   pathological   anatomy.    153 

strawberry,   342 

stiychnine.   390,    392 

venom,   loss   of  bacterial  power,   155 
thrombi    from,    153 

viper,  152 
Poisonous  bacterial  proteins,   131-132 

fish,  162-164 
Polished  rice.  287 

Pollen,   active   toxic   constituent  an    al- 
bumin,   147 
Polyamino-acids,    285 
Polycythemia,  293,  313 
Polymerization,   429,   638 
Polyneuritis  gallinarum,  287 
Polypeptid,  20.  282 

synthetic,    194 
Polvphenoloxidases,    71 
Polyplasmia,   303 
Polypoid   tumors'.   428 
Polysaccharides.   656 
Polyuria.    015 
Porges-Heriiiaun-Perutz  reaction.  237- 

238 
Porphyrins.   480 
Portal   vein   thrombosis,  354 
Positive  cheiiiotaxis.  250,  271.  270 
Postmortem  changes.  71,  101-102 

decomposition.  477 

hemolysis.   224 

lymph'  flow.  346 
secretion.   332 
Potassium,  26.  45.  303,  499 

chlorate.  232 

cliloride,   336 

])liosphate.   290 

salts.  48.  257.  284.  525 
Potocytosis.  333 
P-oxyphenyl-lactic    acid.    543 
Pieci])itate,   resistance  of.    190 
Precipitation  of  colloids.  40.  ]S7.  301 

S|iecific,    507 
Precipitinogen.    189 
Precipitins.    179,    189    193.   240 

barterial.    192 


INDEX 


699 


Precipitins,   cluiiiical    jjiopcrties,    193 

for  ilyt's,    ITii 

iuiptopliori',   I'.H 

reactions.    l:i!t.    358 

relation  to  anapliylact in,  li);5 
rrecipitoid,   1!»1 
Precocity,   sexual,    OOS 
Piefrnuncv,   04,   8(),   410,   4:50,   453,   (507, 
012 

acidosis  of,    559 

diagnosis  of,  200 

pernicious   voinitinjr   of,    538 

toxemias  of,  533   539.  .").")7 
Preputial    concretions.    463  464 
Pressor  bases,   125,  507,  576   577 

substance,  007,  014 
Primary   toxicity    of    foreign    sera,    203 
Proliferation,  283   285 

chemistry  of,  2S4 
Propionic  acid,  507 
Prostatic  concretions,  437,  404 
Prota-ron.   25.    100 
Prolamin.   20,    131,    285,   420 

sperm,    177 
Protease.  81-105.  277,  201,  301 

leucocytic,  03,  05,  103 
Protective  ferments.  107 
Protein,  19-23,  33.  335 

bacterial.  270,  2.S0 

poisonous,  131-132 
toxic,  not  specific,  132 
toxicity  of,   131 

Pence- Joiies,   30!),  518,  570 
constitution   of,   520 
reaction  of,  519 

blood,  290 

cell,  autolysis  of,  403 

chanp:es  in,  440 

cleavajre   products,    194 

coagulated,  383 

compound,    21 

concentrated,  420 

concretions,   452 

contents  of  edema  fluids.  356-357 

etTects  of  skin  injection,  200 

electro-negative,  396 

foreign,   165 

hydrolysis,  569 

insoluble,   23 

intoxicating  dose,  195 

katabolism,  increased,  302 

metabolism,  58,  444 
in  l(nd<emia,  309 
in   ])ernicious  anemia.   300 

molecule,  chemical  composition,  20 

natuie  of  enzymes,   54 

of   cell.    21 

origin   of.    521    522 

putrefaction,   571    583 

pyogenetic.    lOS 

pyogenic,  270 

racemized,  of  Dak  in.  100 


Protein,     rciation     of     andniceptor     to, 
214 
of  antitoxins  to,    isl 
of  phytotoxins  to,  144    145 
of    toxins    to,    120 

sensitizing  dose,    195 

simple.  21 

solubility,   ell'ect  of   light  on,  375 

sugar  from.  603 

tumor,  same  as  normal  i)roteins,  494 

vegetal)le,    270 
Proteolysis,  autolytic,  520 

tryptic,  520 
Proteolytic,  enzymes,  280,  315 
of  leiicocytes,  94-96,  277 
Proteoses.   281,   320,   357,   507,   508 

coagulation   inhibition,   508 
Proteroglypha,    149 
Prothrombin,  290,  295.  299,  316 
Protomain    endestinate,    100 
Protoi)lasm,    18,    41-43 

elFect   of  electricity   on,   378 

foam    structure    hypothesis,    51 
Protoplasmic   poison,   379 

streaming  255.  2(iS 
Protozoa.    135-137.   252.    376,   432 

ciliated,  255   256 

immunity  against.  135 

pliagocvtosis  by,  262 
Pruritus,"  487 
Prussian  blue,   290 
Pseudochylous  effusions,  304 
Pseudo-globulin,    ISl,    290,    351,    356 
Pseudoleukemia.    312 
Pseudonielanosis.    481-482 
Pseudomucin,   420,  513 
Pseudomyxoma  peritonei,  514 
Pseudopodia,  200.   272 
Pseudo-rachitic  lione.  440 
Pseudo-rickets.  447 
Psychosis.  301.  500 
Ptomains.  120-125,  3S9.  507 

chemical  composition  of.  122 

lisli,  103 

no  imnumity  against,   122 

non-specificity  of,   121 

poisoning,  sources  of.  122 

relation  of  toxins  to.  128 

resemblance    to    vegetable    alkaloids, 
121 

structure  of,  121 
Phyalin.   salivary,   429 
Pu'beity,   007 
Puerperal  eclampsia,  78,  539 

uterus,   393,   509 
Pulmonary  edema,  281 
acute",  351 

gangrene.   281.   389 

hyperemia.   341 

inllammation.   405 

tub<>rculosis,   79.   407,  505 
ulcerating.  509 

veins,  439 


700 


INDEX 


Purine,    04,   248,    357,   497-498,    584, 
618 
bases.  22,  310.  525 

bodies,  280,  (IIS 

enzymes,  497  498 

excretion,    (il4 

nictaljolisiM.    fi20 

nitrogen   in  tumors,  4!t5 

nuclease,  (522 

nucleus,  618 
Purine-splitting  enzvmes,   105 
Purpura,    2n7 

hemorrhafiica,   230,  2!)7,   317 

neonatorum.    322 
Purul(>nt    tluicls.    358 

pneumotliorax.  3ti5 
Pus,  70,  78,  93-94 

cocci,   129 

composition  of.  277-280 

decomposed,  414 

inspissated  collections  of,  438 

serum.    278   279 
Pus-collections.  ins])issated,  415 
Pus-cor])nselcs,   278 
Pustule,   malignant.   276 
Putrefaction,    122,   388,   410,   570-589 

intestinal,  245,  588 

lack   of,   362 

of  antitoxin,   182 

protein.  571-583 
Putrefactive  bacteria.  566 
Pntrescine.    123,  567.   582,  583 
Putrid  bronchitis.  434 

cancers.  574 

]>lacenta,    574 

pneiunothorax,    365 

purulent  exudates,  574 
Pvcnosis,  07,  285,  328,  369 
Pyin,  279,  421 
Pvocvaneus  toxins,  218 
Pyocyanin,  133 
Pyocyanolysin,    224 
Pyogenetic  proteins,  108 
Pyogenic  ]>roteins.  276 
Pyonephrosis.    433 
Pyopneumothorax,   365 

open.    365 
Pvosalpinx,  451 
Pyridine.  247,  250.  521 
Pyrimidine.  22.  287.  619 
Pvrocatechin.   575.   612 
Pvrogallol.   217.   249 
Pyrrole.   469 

ring.  471 

QUADRU'RATE,    620 

Quinin,  251,  257,  258 

l!Ari:Mi/.i:i)  ])r(itciii   of  Dakin,   l(i6 
Padiating  structure.  448 
Padicals,  aminoacid,   176 

aromatic,   176.   191 

in    molecule,    arrangement    of.    176 


Radium.    60.     104,    501 

necrosis.   376 
Pancid  oils  in  alkaline  solution,  207 
Hauid  elimination,  245 
Hattlesnake   venom,    150,    228 
Rays  serum,  toxicity  of,   164 
Reaction   of  degeneration,   392 

reversibilitv  of,  56 
Real   alkalinity-   291 
Receptaculum   chvli,   303 
Receptors,  129 
Red  blood  cells.  70 
corpuscles.  289 

degeneration.  498 
Reducing  enzymes.  74 
Reduction.  247,  248 

chemical   changes    in   crystalloids  bv, 
33 

of  methylene  blue,  253 
Refractoriness,    2-14 
Regeneration,   253-288.  371 
Renal  disease,  lipoids  in,  609 

dropsy,    344 

edema.   348-350 

elimination,  524 

e])ithelium,  calcification   of.  438 

pelvis,  457 
Rennin,  61,  67.  115.  291 
Repair     and     growth,     chemical    basis, 

285-288 
Respiration,  internal,  548 
Respiratory  metabolism,  444 
Retinitis,   albuminuric,  530 
Reversibility  of  reactions,  56 
Rhabdomyoma.    432.   497 
Rheumatism,  321,  480 
Rheumatoid    arthritis,    629 
Rhinoliths,  459.   464 
Rhi/opods.   262 
Rhubarb,   457,    584 
Rims    diversiloba,    147 

toxicodendron,   147,   167 
Rice,  polished,  287 
Ricin.  75.  144.  177.  223.  225.  293,  451 

agglutinating  poAver,  145 

phytotoxin.  167 

]ioisonintr.  245,  573 

histologic  changes,  146 

toxicity  of,    145 
Rickets,  287,  445-447 

relation   to  osteomalacia,  446 
Rigor,   antemortcm.   390 

mortis.  101.  390  393 

relation  to  acidity.  391 
Robertson  film  theory,  51 
Robin.    144.    225 
Roentgen  rays.     See  also  X-ratts 
Rovida's  hvalin  substance,  279 
Rubber.  36' 
Russell's   fm-hsin  bodies,  424 

SAcruAUosK.  280 
Saccharosuria.  657 


INDEX 


701 


Siifiiuiiii,    I'M) 

JSiifjo  spliH'ii,  4 IS 

Saliiinaiiders,    poisons    in    skin    of,    162 

Salicyl-alcU'liyde,    72 

Saliovlic   atid,    250 

Salivary  calculi,  462 

phyalin,   42!) 
Salmon,  spawn inj,',  420 
Salpin<,ntis.    4:!;? 
Salts,  71,  290,  :332,  -.Ul 

alkaline,  ;3."37 

bile,  487,  500  . 

calcium,  4S,  317 

fractionation  iiictliod.   ISl 

inoi'^'aiiii'.  2S,   4S,   .")  1 

j)otassiuni.  4S,   32") 

relation   of,  to  ag<:lut  inal  ion,    ISO 
Salvarsaii,  243 

anapiiylactic    reactions    with,    Iti!) 
Samanilaridin,    102 
Saniandarin,    lti2 
Sand,  intestinal.  4(i;! 
Saponin.  217.  226   228.  244 

etrect  of,  227 
Sajionin-cJiolesterol    coinpound.   220 
Sajjonin-lecitliin    compound,   220 
Sapotoxin,  227 
.Sapremia,  380 
Sareoeystin,   137 
Sarcolaetic  acid,  543,  555-557,  501 

relation  of  oxygen  to,  555 
Sarcoma,  437,  468,"  497,  634 

spindle-cell.  494 
Sarcosporidia,   135.   137 
Saturation  limit,  ()39 
Scarlatinal  virus,  specific.  235 
Scarlet  fever,  252,  321,  349 
Sears,  corpus  luteum,  480 

episplenitis.  424 

tissue,    424 
Sciatica,  586 

Sclercnchymatous  ^articles.  463 
Sclerosis,  multiple.  414 

senile.  439 
Selerostoma  equinuni.  143 
Scorpion  poison,   157-158 

toxin,   177 
Scorpcena   scor]ilia.   162 
Scurvy,  286.  297 

infantile,  287 
Sea-snake  venom,  155 
Sea-urchins.   2S4 

eggs,  47.  70 

developing.  2S5 
Sebaceous  glands,  465 

material.  515 
Secondary  anemia,  225.  294,  300-302, 

317,    322 
Secretory  activity,   increased,   343 
grannies.   49 

theory  of  Ivnipli   formation.  332 
Selenium.  247 
Self-digestion  of  liver,  75 
Seminal   vesicles,   474 


Semip('iiiicai)if    mcniliraMes,    30 
Senile  osteoporosis,   444 

sclerosis.  439 
Senility,  371,  5S7 
Sensitization,    187 
active,    191,  202 
by  brilliant  green,  220 
by  silicic  acid,  220 
passive,  191,  203 
Sepia  from  squid,  468 
Sepsis,  350 

local,   433 
Sej)tic  conditions.  485,  610 
infections.  7  1 
softening,  329 
Septicemia.   224.   319,   433.   5.39 
Septum,  nasal,  423 
Serine,  495 
Serosamucin,  357 
Serozyme.  31(i 
Serum    albumin.   21.   29() 
antiplatelet,  239.  300 
anti|U()tense,  277 
bacteriolysis,  208,  210-214 
Idood.  29*0 

colloids,    electro-positive.    452 
eel,    177,   229,   318 
poisons  in.    164 
endotiieliolytic,  239 
foreign.    322 
globulin.  205 
iieniolysis,  mode  of.  222 
influence  of.  (m  phagocytosis.  264 
Lami>rev,  toxicit\'  of.   164 
leucocvtolvtic.    239 
lymph'atolytic,  240 
myelotoxic,  240 
neurolvtic,  240-241 
l)us,   278   279 
Rays,  toxicity  of.  1()4 
reaction.   Klausner's.   237 
snake,    156 
therapy.    130 
thyrolytic.   241 

treatnient     of     e\i>phthalmic     goiter, 
605 
Sex  cells,   174 
Sexual  inactivity.  614 
precocity.  608 
und(>velopment.   614 
Sheep  spermatozoa.  173 
Shellfish    jioisoning,   342 
Shells  of  invertebrates.  437 
Shock,  ana])hylactic.   320 
Sialolithiasis."  4ti2 
Siderosis,  46(5 
Silicates.  465 
Silicic  acid.  34.  40,  223 

sensitization  l»y,  220 
Silver.  246 

nitrate.   276.   294 
Sistrurus.    149 

Skatole.    248.    258,    469,    567.    572,    575 
Skatole-carbonic  acid,  567 


702 


INDEX 


Skatoxyl,  240.  572 

Skeletal  growth,  591 

Skepto-phylaxis,  199 

Skill  cancels  in  colored  races,  467 

changes  in,  614 
Smallpox,  252,  570 
Smegma,  463 

Snake  poisoning,  patliological  anatomv, 
153 

serum.  156 

venom,  148-157,  177,  223,  293,  318, 
319,  564 
Snake-bites,  mortality  from.   151-152 
Soaps,  25,  65,  102,  279,  4U7 

calcium,  387 

formation  of,  441-442 

cysts,   515 

of  fatty  acids,  384 

toxicity  of,  414 
Soda  dropsy.  350 
Sodium  bicarbonate,  218 

diphosphate,  455 

glvcocholate,   237 

salts,   257 
Solanacese,  223 
Solanidin,   227 
Solanin,  227 
Solidago,  147 
Sols,  34 

Solubility  of  colloids,  36 
Solution  of  intercellular  substance,  343 

tension.    187 

true,  of  crystalloids.  35 
Spawning  salmon,   420 
Species  specificity,   176 
Specific    antibodies,    therapeutic    stimu- 
lation  of,   174 

precipitation,  507 

scarlatinal  virus,  235 
Specificity   altered  by   cliemical   charac- 
teristics.   17(i 
by    physical    measures,    176 

its  dependence  of  biological  relations, 
171    172 
on  chemical  compusitioii,  172-173 

of  anti-cnzynies,  M'> 

physico-chemical  factors.  175 

species.    176 
Sperm   histones,    177 

prot.amines,    177 
Spermatocele  fiuid,  359 
Spermatotoxin,   241 
Spermatozoa.   62,    214.    23S.   256,   285 

heads  of.  44 

shec]),    173 
Spermatozoids,   254 
Sperm  in.   2S0 

crystals.  311 
Sphcroliths,  452 
Spideis,   60 

poison,    158-159 

toxin,   177 
Spina   bifida.   360 
Si)inach,   457,   584 


Spindle-cell  sarcoma,  494 
Spleen,  296 

autolysis,    microscopical    and    chemi- 
cal   changes,    369-370 

sago,  418 
Splenectomy,   229.   232 
Splenic  amyloid,  420 

hemolysis,  232 

infarcts,   328 

tumor,  486 
Splenolysin,   241 
Splitting  olT  of  water,  248 
Spores,    112 

bacterial.  367 
Sputum,  93,  96,  280-283,  433 

asthmatic.   311 

bronchiectatic,    4 1 4 

chemistry  of,  280   283 

pneumonic,  490 
Squamous  cell  cancer,  516 
Stagnation,    323,    452 
Staining      bacteria.      Gram's      method, 
112-113 

fat,  in  tubercle  bacillus.  111 

properties  of  amyloid,  420—421 

reactions  of   bacteria,    112-113 
Stains,    fat,    401 

vital,  50 

permeability  for,  371 
Stalagmometer,    208 
Staphvlococcus,  70,  78,  92,  94,  224,  256, 
278 

aureus,   320 

pyogenes,  264,  276 
aureus,    114,   132,  475 
citreiis,    132 
Staphylokimase,    117 
Staphylolysin.  224 

characteristics  of,  224 
Starch,  34 
Starvation,   292,   302,   393,   394,   559 

relation  to  autolysis,  87 
Stasis,   314 

general,    341 
Stearic  acid.   514 
Stearyloleyl    lecithin,   24 
Steatogenetic  poisons,  403,  409 
Steatosis,  cholesterol,  405 
Sterilized    bacterial    cultures,    258 
Stimulation,  mechanical,  263 

tactile,    262 
Sting  of  insects,  351 
Stomach,  cancer  of,   104.  504 

effect  of  venom   in,   152 
Stone.     See  Cdlciiliis 
Strangulati'd    hernia.    341 
Strawberry  juice,  332 

])oisoiiing,   342 
Strci)t<icoccus,    92,    94,    289,    320,    451 

viridaiis,    225,    482 
Strci)t()<-olysin,  224 
Streptothrix.    168 
Stronlium,  444 
Struvit  stone,  457 


IXDIJX 


703 


Stiycliiiiii,   2.")1.   ii.')2 

poison iiij;,  3!tU,  3'.)2 
.Sul)CiitaiK'oiis   ollusioiis.   35t) 
Suhstaiici's  t'l'i'lily  chi'motac-tic,  2o7 

ncjiutivi'ly  clu'motactic.  2')1 

\villi  strong  positive  ciieniotaxis,  258 
Succinic  acid,   507 
JSuccus  cntciicus,   02 
.siijjar,  :vA-2.  :\:,: 

lilood.  641 

concentration.    041 

content  of  blood,  o.'Jo 

from  fat,  00.3 

from   otlicr   substances,    004 

from  protein.  003 

state  of.  in  blood,  643 
in  cells,  044 

supply  to  kidneys,  637 

tolerance,   613 

utilization    of,    038 
Sugar-splitting   enzymes,   55 
Sulpliemoglobin,  481,  582 
SuJpliemoglobineniia.    580 
Sulphides.  247 

metallic.  247 
Sulphocvanides,   251 
Sulphon'al,  480 
Sulphonic  acid  dyes,  50 
Sulphur.  251,  468 

cystine,   oxidation   of,   014 

elimination,  506.  534 
Sulphur-niethcmoglobin,   481 
Sulphuric  acid.  248,  336 
Suppuration,    66,    92,    119,    276-280, 
319,  382,  569 

inhibited   by   excess   of   serum,    277 
Surface    configuration,    1 75 

forces.    175 

phenomena,   56 

tension,  39,  187.  267,  355,  301 

explanation,  objections.   274-276 
Suspensions,  35 
Sweat.   490 

Swellinsr,  cloudy,  394-396 
Sympathetic   nervous   system,   472,   563 
Svnanceia    l)rachio,    103 
Syncytiolysin.  241,  536 
Svncvtioma,  497 
Syncytium,  393 
Synovial    fluid,   427 

mcndjranes,   pathological.   514 
Synthesis   by   enzymes,    57 

in  body,   250 
Syntiietie  activity  of  bacteria.   109 

polypeptid.   194 
Svntonin,  392 
Svplulis,    79,    229.    230,    233.    3()0.    422. 

542 
Syphiliiic  lesions,   407 

TAnEs.  231 

Tabetic  arthropathy.  351 
Tactile  stimulation,  202 
Tienia,   139    140 


Ta-nia  ecliiMococeus,   137 

margiiuita,   138 

perfoliata,    140 

plicata,  140 

sagiiiata,    138,    139 
Tannin.  34,  249 

Tape  worm  lipoids,  antigen  from,  108 
Tar,  anthracene  fractions  of,  493 
Tarantula,    159 
Tellurium,    247 

Temperature,  bi;dy.  sui)nornial,  015 
Teratoma,  432.  497 
'ler penes,  21S 
Testicles,    474 

x-ray  atrophy.  ;>70 
Testicular    tumors.    432 
Tetanization.    344 
Tetanolvsin,  224 
Tetanus,  390 

bacillus.   70,   177 

toxin.    70,    75,     103,     128,    260,    379, 
592 
digestion  of,  127 
Tetany,  287,  581!,  587 

parathyreopriva,  598 

parathyroid.  549 
Tethelin,    015 
Tetrodo-toxin.   104 
Tetrodon,  164 

poison    in    ovaries   and    eggs,    103 
Tetroses,  ()48 
Theobromine.  248,  019 
Theophyllin,   022 
Therapeutic  immunizations,   173 
Thermal   injuries.   253 
Tliermic  alterations,  372-373 
Tliermoprecipitins,    191 
Thermostable  antilipase,  79 
Thermotactic  edect,  255 
Thermotaxis.  2()9 

of    leucocytes,   261-262 
Thermotro])ism.    255 
Thigmotropism.   25(1 
Thoracic  duct,  occlusion  of.  340 

lymph,  osmotic  pressure  of.  33!> 
Thorium.  21 1 
Thorium-J".  259 

action  on  leiu'ocytes,  377 
Thrombin,  291.  316 
Thrombogen.   310 

Thromhokinase,    299,    310,    320.    380 
Thrond)oplastin.  310 
Thrombosis,    105.    303,    315-325,    340 

tibrin-ferment,  325 

portal  vein,  354 
Throndius.  223.  293 

agglutiiuitive.    325 

bile.    485 

librinous.  323 

formation  of.  322-325 

hyalin.  224.  324   325.  374 

secoiulary  clianges   in,   325 

tvphoid."224 
Thvmine.  109.  019 


704 


INDEX 


Thviiiol.  24!1 

Tliyiims.  4!t4.  616   617 

extirpation  of,  (iHi 

liiston,  4 lit 

mick'ic  ai-i<l,  021 

tissue,  retrogressive,  404 
Thyreoglobulin.    5J)3,    (100 
Thyroid,  cancer  of,  (504 

catalase.   5!)") 

chemistry  of.  593-597 

colloid,   426.    .">!*;! 

cystic,    42(1 

detoxicatoi'X'    fuiictidii    of,   .")!)! 

diseases  of!  590   593 

effect    on     carhnln  drate     iiietai)olisni, 
r)!)l 
on  growth.  .'Jlll 
on    liver    autolysis.    .191 

extract,    therapeutic   use,    G02 

functions  of,   590-593 

lipase.  o05 

peroxidase,  595 

relation  to  metabolism.  590-591 

secretion,  deficient,    590-591 

tissue  loss  of,  590 
Thvroidectomv.    241.    002 

effects    of.    591 
Tlivroidisnuis.    004 
Tliyroiodin,   128.  420.  593 
Tliyrolytic  serum,  241 
Tissue  coagulins,  323 

cultures,    40S 

disintegration,    258 

effect  of  light  on.  374-377 

necrotic,   97 

relative  susceptibility  to  heat,  373 
Tissue-cells,    autolytic   enzymes   of,    277 

beluivior  of.   273 
Toad  poison,  150,  161-162 

in    blood,    101 
Tolerance,   acquired,   246 
Tonsillar  concretions,  4()5 
Tonsillitis,    433 
Tophi,   gouty,    631 
Toxalbumins,    144 

bacterial,    108 

vegetable,  225 
Toxemia  of   pregnancy,   533-539,   557 
Toxic    bacterial    proteins,    not    specific, 

132 
Toxicity  of  acetone,  553 

of  ascaris,  141 

of  en/viiies.  61    63 

..f  indole.   574   575 

of  ui'ine.  534 
'{"oxin-antitoxin.    180 
Toxins.   120,  125    129.  177.  394 

and  antitoxin,  dilfusion  of,    1S2 
niterabilily  of.   182 

adaorj)tion   of,   127 

agencies     destroving     or     modifving. 
127 

bacterial,  125,  105,  265,  318,  407 

chenucal   ])ii)])ertie8  of,   125 


Toxins.    di]jhtheria,    75,    87,    102,    103, 
127,  128,  260,  317 
digestion    of,    127 
etrect   of    acids,    bases,    and    salts   on, 
127 
of  light  on,  375 
of  j'-rays  on,  127 
Elirlich's  conception  of  nature  of,  128 

theory  of,    177-178 
fatigue,  75 

immunity  against,  128 
neutialization   of,  bv   antitoxins,    170 
of  fatigue,   561-562 
relation    to    <'nzymes,    126 
to    proteins,     126 
to  ptomai'ns,    128 
resemijlaiice  to  enzymes,  67,  68 
scorpion,   177 

specific,  synthetic  products,  128 
spider,    177 
susceptiliilitv   to.    128 
tetanus,   70,'  75,   103,    128,   260,   379, 
592 
digestion   of,    127 
Toxoid,  120 

Toxolecithidin  in  bee  poison,  100 
Toxophore.  07,  128,  178,  212 

grouj),    120 
Trachinis  draco,   162 
Transudates,    94,    331 

nephritic,  composition  of,  354 
Trichinella,  chemistry  of,  142 

infection,  complement  fixation  of,  142 
spiralis,   134 
Trichinosis,  intoxication  of,  142 
Trimetliylamine,   258 
Trional.   480 
Trioses.  645-649 
Triton  ta?niatus.  262 
Tropisms,  theory  of,  255-256 
True  solutions  of  crystalloids,  35 
Trypanosomes.  130 
tolerance  of,  252 
Trypsin,   62,  05,  07,   103.    128,  358.  566 
Trvpsinogen,      activation      by      kinase, 

221 
Trvi)tic  proteolvsis,  526 
Tryptophane,   195.  286.  469-471,  571, 

574 
Tubercle  bacillus.   70,   79,  99,   107,   131, 
132,  204,  320 
antigen   from,   108 
composilioii    of,    108 
fat   stainiiiLT   in.    Ill 
fats  of,    110   111 
fatty  acids  in,  1 12 
modification  of  acid  fastness  of,  111 
no  cholesterol   in,   111 
])lios])hati(ls    in.    1  1 1 
Tubercles.   382,   431 
calcified,  464 
caseous,  27() 
Tiibercnlin,  87,   103,   130 
injections,   260 


INDEX 


705 


Tulh'iculin  rcai't ioii,  570 

witli  (Icntt'io-alliuiiKisc,   17  t 
'I'liliiTculonastiii,    Ids 
'rulicifuldMamin,  .SS.'{ 
Tubi'r<ul()sis,    (Ki,    7S,    !t2,    !»4,    !tS,    27.'{. 
•S'rS,  ;}.')."),  381,  417,  480,  5!).j, 
5!)(i 

autolysis   of   livers   in,    101 

hacillus   of.      St'c   Tubercle   lincillus 

fclirilc  cases,   282 

lviii])li(>c\  tes    of,    2()0 

piiliiioiiaiy,  7!t.  407,  50,5 
uleeratiiifj,  50!) 
'riil)eicui<)iis  aliseess,  cold,  280 

etl'usion,  353,  433 

exudates,  358.  304 

lueuiujritis.  300 

pleurisy    350 

])roiesses,  340 
'J'iil)i)-()varian   cysts,   514 
Tuu^^stcu.    240  ' 

'luiiiors,      104-105,      311.     340,     405, 
422 

adrenal,    melanotic,    472 

autolysis    of,    4!)() 

l)cnij:n,  chemistry  of.  509   515 

hone-marrow,  570 

cells.  404 

chemistry  of.  492-522 

cholesteatomatous,  415 

colloidal  poisons  in,  503 

efVect  of  diet  on  growth,  493 

embryonic  orifjin,  4!)(i 

en/ymes  in,  75 

ferments   in,    105 

•rlycoiren    in,    431-432 

hemolytic  sulistanccs  in.  504   505 

inorganic      constituents      of.      499- 
500 

internal  secretio!i.  502-504 

iron   in.  500 

malignant,  284,  5(;!» 

chemistry  of.  515   518 

melanotic.  '4G7,   471   472,  496,  502, 
577 

mouse.  173.  432 

of  brain.   531 

ovarian.  420 

oxidase  in.  70 

paratlivi-oid,   432 

polypoid,    428 

proteins  of.  same  as  normal  jtroteins, 
494 

purine  nitrogen  in,  495 

splenic.  480 

testicular,  432 
Turgor  of  plant   cells,   30 
Tur])entine  exudates.   358 

oil  of.  270 
Tvi)lioid  agtrlutinin,  174.  184 

bacillus,   70.    129.    132.    183.   211.  214. 
224.  204.  320,  451 

colon   bacteria,   dilTerentiated   by   acid 
agglutination,  188 


■lyi'lKiid      fever,     71,      119,     293,     321, 
.{50 
niin-specirK-  reactions  in,   174 
inlectidii,  .'JOS 
tlirnnibi,   224 
'lypholysin,  -224 
Tyramine,  570 

Tyrosinase,  00,  07.  71,  73.  409,  545 
Tyrosine,  20.  87,  !)0.  258,  279,  309,  .327, 
357.   389.   4()!»,  542,  507,  57S, 
009 
Tyrotoxicon,  585 

Ui.CKRATixo      pulmonarv      tuberculosis, 

509 
I'lceration,  508 
ntra-violet  rays.  00.  182,  375 
I'ncinaria,    318 

duodenalis.    142-143 
Unicellular  organisms,  252 
I'racil,   109,  019 
Uranium  nephritis,  344 
Lrate.  454 
bibasic.    020 
calculi,  456-457 
monobasic,  ()20 
nionosodium.  020,  029 
Urea,  53,  250,  251,  258,   332,  357,  525, 

626,  627 
Urease,  67 

Uremia,  70,  417.  525-533.  534,  560 
acids  in,  528 
asthenic,  529,  532 
chemical  changes  in,  527-529 
etiolofry  nf.  529-533 
relation   of  eclampsia   to,  533 
Ureu'ic  coma,  355 
endocarditis,  531 
pericarditis,  531 
Urethra,  457 

Uric  acid    75,  103,  357,  525,  529,  619 
calculi,  455  456.  027 
chemistry  of.  618  619 
concretions,  310 
deposition,    029 
destruction  of,  625-626 
diathesis,  027 
endogenous.  010,  021 
exogenous.  (i21 

increased   elimination    of.   456 
infarcts.  450.  027.  633-634 
intoxication,  027 
metabolism.   618   634 
oxidation,  248 
properties   of,    620-621 
i-eteiition,  029 

sympathetic  formation.  624-625 
Uric-acidemia.  032 
T'ricase.   71.   95,  623 
Uricemia.    627,    631 
Uricolvsis,   625 
Uricolyfic    en/ymes.    501,   633 
T'rinarv  bladder.  42.3 
calculi.  454  460 


706 


INDEX 


Urinary  calculi,  disintegration  of,  459- 
460 
general  properties,  459-460 

changes,    HXi 

constituents,  toxicity  of,  52.5 

dextrin,  3SS 

pigments,    525 

secretion,  .'i.Sl 

toxiiit\,   525 
Urine,  (i4,  542-544 

amylase  in.   SO 

catalase    in,    71 

concentration   of,  45(J 

fat  in,  320 

lipase  in,  78 

toxicity  of,  53-4 
Urinod,  530 
Urobilin,   230,   2!)U,   302,   450,   474,  477, 

4!)0 
Urobilinogen,  477 
Urobilinogenuria.    4'.I0 
Urochronie,  450 
Ura?rytlirin,   450 
Uro-fuscin,  480 
Uroleucie   acid,  578 
Uronielanin.   450 
I'rorosein,    575 
Urostealitli  calculi.  458 
Urticaria,  351 

factitia.  380 

local,   342 
Uterine  fibroid,  510 

degenerating.  4!)8 
Uterus,   involution   of,   02 

puerperal,  303,  50!) 
Utilization  of  sugar,  038 


Vacuomzao'ion,  378 
A'alerianic  acid,  507,  583 
Valvular  heart  disease.  355 
Vaquez-Osler   disease.   313 
Avascular  disturbances,  253 
Vasoconstrictor  nerves,  253 

paralysis,  312 
Vasoconstrictors.   488 
Vaso-depressor,    GOO 
Vasodilator  nerves,  253 

stinuihis,  312 

stimulation,  351 
Vasodilatoi's,   488 

Vegetalile  ])ois(>iis.  heniolvsis  by,  225- 
226 

proteins,  270 

toxall)umins,  225 
Veins,  pulmonary,  430 
Venom.   105.  245 

agglutinin,    154 

as  antigens,   150 

chemicnl   constitution  of,   150 

cobra.   150.  151,  227,  228,  241 
resistance  to,  505 

<(il)pcrhead,  228 

crotiiJMs,   151 


Venom,  ell'ect  of,  in  stomach,  152 
on  bluod,  153 

enzymes  in,  150 

gland,    140 

hemolysis  by,  228   229 

immune  serum,   320 

Krait,    155 

nu)cassin,   150,  228 

nature  of,    153-155 

ni'urotoxin,  154 

platypus,  157 

poisoning,  loss  of  bactericidal  power, 
155 
thrombi    from,    153 

properties  of,   149 

rattlesnake,   150,  228 

snake,    148-157,   177,  223,  203,  318, 
310,  504 

toxicity  of,   151 

variations  in,  155 
Veronal,  480 

Vertebrates,  cold-blooded,  105 
Vesicants,  351 
Vessel  injury,  323 
Vinegar  eels,  370 
Viper  poisoning,   152 
Viperida^   148,   149 
Viscosity.    203 

of  blood,  292-293.  301 
Vital  activity,  53,  333,  338 

stains,  50 
Vitalistic  school,  330 
Vitamines,  285-288 
Volatile  fatty  acids.  280 
^'omiting,   245,   5()3 

cyclic,  557,   559-560 

hysterical,  500 

pernicious,  of   pregnancy,   538 

Wasps,  100 

Wassermann  reaction,  108,  234-238 

Water  aliinity,  371 

amount  in  Ijlood,  331 

caj)acity  of  colloids  for.  336-337 

dielectric   constant,   38 

distilled,  257 
Waxy  degeneration.  3(i0.  370 

of  muscles.   392   393 
Wet  brain,  531 
White  chromogen.  400 

kidneys,   large,   405 

wool,  400 
Wool,   white,   400 
Worms,  intestinal.  03,  432 

nu-al,   400 
Woniid    secretions,    302 

Xantiiki.asma.  415 

nuiltiplex,  47(i 
Xanthine,  75,  247,  454.  Glfl,  023 

bases.    101 

iiodies,   010 

calculi,  458 

oxidase,  71,  408,  501 


INDEX 


707 


-Xaiitlioiiiii,  4().>,  4SS 

tiilifitisiMii   imilliplc.v,   r)12 
Xiiiit lioiiiatoiis   iiiassi's,   41(i 
Xaii11i(>iiliyll,   47.') 
Xaiit liosiiu*.  &2-i 
Xaiitliosiiu'-Iiydrolasc.   i'fl'.i 
Xerosis  bacillus,  ."{'JO 
A'-rays,  (iO,  M\ 

atr()])liy  of  ovaries  from.  ."J"!! 
of  testieles  from,   ."^7(1 

cancer  from,  .■?77 

eilect  of.  on  toxins,  127 

jjaiifirene  from,  .■?7ti 

leukemia  from,  .■>77 

necrosis  due  to,  376 


A'-rays  treatiiicnl,   lo:{,  5G9 
Xylose,  4i)7,  04!) 

Yea.st  extracts,  287 
tolerance  of,  252 

Zeix,  194,  280 

Zenker's  waxv  defeneration,  33G 

Zinc,  223 

Zofiprecipitins.   I'.lii 

Zootoxins.   148-164,  203 

Zyiiioi^cn,  ()() 

Zynioids,  (i3,  (17 

Zymophore  group,  120 

Zymoplastic  substance,  299,  31G 


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BIOLOGY 

LIBRARY 

APR  "s  1941 

NiftY  1  2  1941 

AH<     J  1942 

1 

V      W3 

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»..■'("                     ■  '4  '4 

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•^^0801 


6'OLOG 
UBRAR 


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UNIVERSITY  OF  CALIFORNIA  LIBRARY 


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